oa Quantification of the Role of Glycosaminoglycans on the Tensile and Viscoelastic Properties of the Aortic Valve Leaflet

The spongiosa is the middle of three layers of the aortic valve leaflet and contains the proteoglycan (PG) versican and glycosaminoglycan (GAG) hyaluronan (HA) in large quantities. The resulting versican-HA aggregates allow the layer to retain large quantities of water, giving the layer a gel-like consistency. This layer is the least understood layer of the leaflet as it pertains to its role in valvular function and mechanics. However, the GAGs in this layer are perceived to be important to valve function, as the loss of GAGs in bioprosthetic heart valves coincides with its failure. The functions attributed to these GAGs include: (1) lubricating shear between the outer layers during flexure and tension; and (2) filling large volumes, resisting compression, and dampening shock from valve closure. The lack of understanding of GAGs role in valvular function is due to the difficulty in its isolation from the rest of the leaflet. For our study, rather than isolate the spongiosa, we enzymatically digest GAGs from the spongiosa via hyaluronidase. Previous studies have shown that the complete enzymatic removal of GAGs from heart valve leaflets greatly increases flexural rigidity. In this study, we instead varied the amounts of GAGs in the leaflets and investigated the effects on the tensile properties of the native leaflet; native leaflets in tension have a bilinear stress-strain curve, little hysteresis and minimal relaxation. GAGs were depleted using varying concentrations (0, 1, 2, 5 U/mL) and application times (8, 24 h) to yield a gradient of 4 GAG amounts ranging between 20-40 μg GAG/mg dry weight of native tissue, although with little change in water content. GAG depletion from the spongiosa was also confirmed using Alcian blue staining, which also verified no gross changes to the outer layers. Leaflets in these GAG depletion gradient conditions are in the progress of being mechanically tested to elucidate the effects of GAGs on the tensile elastic and viscoelastic properties of the native leaflet. This work is a component of a broader effort to elucidate the important role of GAGs and PGs in the function of native and diseased valves and in the design of improved valve replacements.