Scanning Electrochemical Microscopy as a Local Probe of Oxygen Permeability in Cartilage

Abstract

The use of scanning electrochemical microscopy, a high-resolution chemical imaging technique, to probe the distribution and mobility of solutes in articular cartilage is described. In this application, a mobile ultramicroelectrode is positioned close ( approximately 1 microm) to the cartilage sample surface, which has been equilibrated in a bathing solution containing the solute of interest. The solute is electrolyzed at a diffusion-limited rate, and the current response measured as the ultramicroelectrode is scanned across the sample surface. The topography of the samples was determined using Ru(CN)(6)(4-), a solute to which the cartilage matrix was impermeable. This revealed a number of pit-like depressions corresponding to the distribution of chondrocytes, which were also observed by atomic force and light microscopy. Subsequent imaging of the same area of the cartilage sample for the diffusion-limited reduction of oxygen indicated enhanced, but heterogeneous, permeability of oxygen across the cartilage surface. In particular, areas of high permeability were observed in the cellular and pericellular regions. This is the first time that inhomogeneities in the permeability of cartilage toward simple solutes, such as oxygen, have been observed on a micrometer scale.

Viability of equine articular chondrocytes in alginate beads exposed to different oxygen tensions

“…For these authors, 10% O 2 would be present at the surface of the cartilage, with variations of O 2 tension depending on the depth of cartilage (<1% in the deep layers). This estimation has to be re-evaluated on basis of the results of Gonsalves et al (2000), who proved different diffusion gradients depending on the density of the network-like collagen fibres.…”

Oxygen consumption of equine articular chondrocytes: Influence of applied oxygen tension and glucose concentration during culture

“…This lack of O 2 seems partially compensated by a heterogenous distribution: the composition of the cartilage directs the passage of O 2 near to the cells, so that the highest O 2 permeability is in the cellular and pericellular regions (Gonsalves et al, 2000). Moreover, in the deepest layers of the articular cartilage, the availability of O 2 will depend not only on the density of chondrocytes, which constitute just about 1-2% of the equine articular cartilage (Todhunter and Lust, 1992) and on their particular cellular respiration rate, but also on the diffusion and the gradient of O 2 .…”

Quantitative visualization of passive transport across bilayer lipid membranes

“…Dynamic electrochemistry in the form of scanning electrochemical microscopy (SECM) has found some application in the study of membrane transport (31)(32)(33)(34)(35)(36). Our method uses an ultramicroelectrode (UME) placed close to one side of a BLM to deliver weak acids in a well defined manner by the electrogeneration of protons in the presence of weak acid anions and a trace quantity of the pH-sensitive fluorophore, fluorescein.…”