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Cartilage Repair: Effects of mechanical loading
R Lane Smith, PhD; David J Schurman, MD; Dennis R Carter, PhD; B Donlon, BS; BE Ellison, BA; M Gupta, BS; M Mohtai, MD
Effects of Hydrostatic Pressure
Our studies tested the hypothesis that mechanical loading, applied at levels of a magnitude approximating forces generated during normal joint activity, would modulate cartilage extracellular matrix synthesis.
Hydrostatic pressure was applied to high density chondrocyte cultures for 4 hours using computer controlled servo-hydraulics. mRNAs were identified by northern blotting and quantified by slot blots and densitometry. GAG synthesis was quantified by 35SO4 uptake; matrix deposition was determined by immunohistochemistry.
Under these conditions, intermittent hydrostatic pressure (IHP) increased chondrocyte mRNA signals for aggrecan by 7% (p<0.05) in serum-free medium and by 15% (p<0.05) in medium containing 1% fetal bovine serum (FBS). Intermittent hydrostatic pressure increased type II collagen mRNA signal by 15% (p<0.05) in the presence of 1% FBS. Intermittent and constant hydrostatic pressure stimulated glycosaminoglycan (GAG) synthesis by 59% (p<0.001) and 35% (p<0.05), respectively, in serum-free medium. Extracellular matrix deposition was enhanced by intermittent and constant hydrostatic pressure in serum-free and serum containing cultures as determined by immunofluorescence using antibodies specific for aggrecan and type II collagen.
IHP also stimulated GAG synthesis in full thickness cartilage explants by 37% (p=0.06, n=4) at 12 hours of exposure, 32% (p<0.05, n=5) at 24 hours, 55% (p<0.01, n=5) at 48 hours, and 64% (p<0.01, n=5) at 72 hours. GAG synthesis in cartilage control explants increased for the control values by 1.2 fold, 2.7 fold and 3.3 fold at 24, 48, and 72 hour over that of the 12 hour time point. With IHP GAG synthesis was stimulated by 1.0 fold, 4.1 fold and 6.4 fold increases at the same 24, 48 and 72 hour time intervals. All explants were maintained in serum free medium. The results are shown graphically in Figure 1.
These data demonstrate that intermittent and constant hydrostatic pressure stimulate adult articular chondrocyte metabolism and suggest that high levels of hydrostatic pressure in the joint may contribute to cartilage repair.
Figure 1. Cartilage explant cultures with and without exposure to IHP.
Effects of Fluid-Induced Shear Stress
Our studies also tested the effects of deviatoric (shear) stress on adult articular chondrocyte metabolism in vitro. High density monolayer cultures of normal bovine and human chondrocytes were exposed to continuous fluid flow-induced shear stress (1 Pascal) for varying periods up to 72 hours. Under culture conditions in which the cells were distorted by the fluid- induced shear, articular chondrocytes elongated and aligned in the direction of applied stress within 48 hours of treatment.
Shear stress stimulated glycosaminoglycan (GAG) synthesis (50-80%, p<0.05) and increased the hydrodynamic size of proteoglycan monomers, in part by lengthening the GAG side chains. Prostaglandin E2 release into the culture medium was elevated 12-fold; IL-6 was significantly elevated in the culture medium following shear stress (Figure 2). In addition mRNA levels for tissue inhibitor of metalloprotease (TIMP-1) was increased 8-fold.
Figure 2. IL-6 release in response to shear stress.
These data demonstrate that deviatoric (or shear) stress directly influences chondrocyte metabolism and suggests that shear stress may influence articular cartilage homeostasis in vivo.
These studies address, at a molecular level, the basic mechanisms by which mechanical loading may influence articular cartilage metabolism. The long term goal of this work is to develop fundamental knowledge that will contribute to connective tissue repair.
Republished from the 1994 Rehabilitation R&D Center Progress Report. For
current information about this project, contact
R Lane Smith.
