To examine whether differences in the pericellular microenvironment of different chondron preparations influence the chondrocyte volume regulatory response to experimental osmotic challenge.
Mechanically extracted chondrons (MC), enzymatically extracted chondrons (EC) and isolated chondrocytes (IC) were seeded into agarose and sampled at 1, 3 and 7 days. Samples mounted in a perfusion chamber were subjected to osmotic challenge. The cross-sectional areas of the chondrocyte and pericellular microenvironment were measured under isotonic, hypertonic and hypotonic conditions, and percentage change calculated. Separate samples were immunolabeled for type VI collagen and keratan sulfate.
Initially, the microenvironment of MC represented 60% of the chondron area and was occupied by type VI collagen and keratan sulfate. In EC, the microenvironment comprised 18% of the chondron area with narrow bands of type VI collagen and keratan sulfate. IC had no visible microenvironment, with small amounts of type VI collagen and keratan sulfate present. All preparations sequestered additional pericellular macromolecules during culture. Under isotonic conditions, the EC and IC chondrocytes were larger than those of MC. All chondrocytes shrank under hypertonic conditions and swelled under hypotonic conditions. MC were the least responsive, displaying the most efficient volume regulation. IC showed the largest response initially but this decreased with time. EC exhibited intermediate responses that decreased as the microenvironment increased in size.
The composition and structural integrity of the pericellular microenvironment do influence the cellular response to experimental osmotic challenge. This suggests that the microenvironment functions in situ to mediate the chondrocyte response to physicochemical changes associated with joint loading.