Topic 9: Testing Collagenous Tissues

Collagenous Tissues: Summary of Key Points Collagen is a ubiquitous structural protein with many types all having a triple helix structure that is cross-linked in a staggered array.triple helix structure Some of the most common collagen types are fibrillar and the collagen can be organized in 1-D, 2-D or 3-D in different tissues to confer different material properties. fibrillar material properties The 1-D hierarchical arrangement of stiff collagen fibers in ligaments and tendons gives these tissues very high tensile stiffness1-D hierarchical arrangement The 2-D arrangement of collagen fibers in tissues such as blood vessels and intestine is often quite wavy or disordered to permit higher strainsblood vessels intestine

Collagenous Tissues: Key Points (continued) Crimping, coiling and waviness of collagen matrix gives the tissue nonlinear properties in tension.Crimping, coiling and waviness Collagen structure in tissues changes with disease and ageing.ageing The hierarchical cardiac collagen matrix organizes cardiac muscle fibers in three- dimensions. Interstitial fibrillar collagen in the heart wall contributes to tissue stiffness during filling.hierarchical cardiac collagen matrixtissue stiffness

Testing Considerations Structural Properties describe the behavior of the actual tissue (e.g bone ligament bone complex) Mechanical properties describe the behavior of the tissue as a general material

Ligament Tensile Testing computer Cross correlation Strain computation

6 DOF Knee Testing Rig

Clamping considerations Device to hold tissue and clamping must be stiffer and stronger than the subject material. Otherwise the stiffness of the device contributes to what you measure. In biological testing this can be quite problematic. If possible, it is important to measure the intrinsic stiffness of the 'jigs' used to hold the tissue. Ligaments—have their own "built in" clamps -- bones. Usually drill holes in bone use steel rods. Do not want to clamp too far away of elongation may include bone deformation. Do not want it too close because may violate insertion (attachment) of ligament to bone. May weaken bone... the not measuring properties of ligament. Tendons only have 1 "natural" clamp Wherever you clamp, have to worry about inhomogeneities and edge effects. Structural properties have less significance.

Measurement of strain Measurement of deformation of biological tissues is nonhomogeneous, i.e. the different regions can deform independently. Therefore, it we use the "clamp to clamp" strain the measurements would be average over the whole region and any slippage in the clamping system would appear to be relevant. Many approaches have been used to measure strain in biological tissues. Because the deformation in a ligament or tendon is uniaxial, the approaches can be more simple. strain gauge--invasive VDA - Video Dimension Analyzer Digital video imaging

Strain Gages Piezo Electric Crystal F V Mercury Hg F F V Thin Film

Strain Tensors: 1D example Cauchy (infinitesimal) LagrangianEulerian

Stress-free state How can we identify the best 'reference' state for the stress and strain calculations? Recall: Lagrangian strains are referred to original lengths Problems: The soft tissues buckle under compression Long toe region makes it difficult to identify transition from compressive to tensile forces Solution: Use a small tare load to repeatably identify the initial state

Hysteresis Loading & unloading curves are different Area between curves represents energy absorbed by material Anelastic Properties Preconditioning Apparent material properties are history dependent Becomes repeatable with multiple cycles (in ligaments and tendons tested in vitro, this occurs between 4-7 cycles)

Viscoelastic properties Stress-Relaxation stress decreases with time but reaches an equilibrium for a step increase in strain Creep Strain gradually increases with time but reaches an equilibrium for a step load

Strain-Rate Effects In many biological tissues, strain rate - or how fast the tissue is loaded - influences the material properties In general increased strain rate results in increased stiffness due to viscous effects Not such a huge effect in ligaments and tendons, but very important in regards to prevention of injury

Age Similar changes occur in collagenous tissues among individuals and most species. There are successive increases in collagen which eventually becomes more organized and cross-linked until skeletal maturity is reached. Results in raised elastic stiffness and strength. Once an animal/person reaches skeletal maturity, the properties begin to deteriorate. Stress, MPa Strain, %

Skeletal Maturity and Relative strength of substance and insertion sites

Advancing Age As a person becomes older, the maximal force their ACL can tolerate decreases, this is has as much to do with changes in geometry as it does changes in material properties

Immobilization MCL is metabolically more active, so as it remodels the tissue it lays down mechanically compromised material. However, the ACL cannot produce new tissue, so it simply atrophies. Immobilization of the knee causes deterioration of the MCL material properties, but not the ACL material properties. If you were to graph the structural properties, you would see changes in both ligaments.

Knee Ligament Injuries MCL/LCL - HEAL ACL/PCL - DON'T HEAL Mechanism of injury? –Turning/cutting... –Valgus stress, –Anterior tibial translation (clipping) –Hyperextension

Ligament Injury How do you study knee injuries? 1.) Clinical Studies interviews always post injury 2.) Cadaver Studies Don't know kinematics of injury Can't study healing 3.) Animal Models How to make repeatable injury –Cut -- poor healing –tear -- good healing –z-cut -- poor healing –This suggests that an increase in stretch is important for proper healing

ACL Healing no matter what type of model, poor repair Why? –Synovial fluid –decrease vascularity –different cell shape/type –ACL fibroblasts look like cells of cartilage (which doesn't heal) –the $10,000, Question!

ACL Repair Surgery –Usually only do surgery on active people ‹30 –For most injuries, replacement is only form of surgical intervention

Autograft Sacrifice another tissue from same person –e.g. 1/3 quad tendon –Pro rejection unlikely readily available –Con decrease function of donor tissue different tissue - different behavior Autograft -- Sacrifice another tissue from same person

Allograft Cadaver donor –Pro no loss of function same species better acceptance –Con rejection disease availability

Prosthetic/Tissue Engineering e.g. Dacron; GoreTex (Tissue Engineered not yet available) –PRO Inexpensive to manufacture No biohazard worries No decreased strength (unless resorbable stent) Less rejection –CON not the same scar formation fatigue/wear Design Characteristics –like ligament (non-linear toe region with transition to a linear phase) –promote biological adaptation –maybe grow fibroblasts first –put matrix for fibroblasts to attach to bio-compatible materials Petrigliano et al. Tissue Engineering for Anterior Cruciate Ligament Reconstruction: A Review of Current Strategies. Arthroscopy 22(4):

Xenograft Donor tissue from another species; e.g. porcine –pro readily available no disease problems no loss of function –con rejection (usually irradiated to lower risk) weak during remodeling (several weeks)

Surgical considerations Pretensioning Joint Kinematics –6-DOF –Displacements Anterior-Posterior Medial-Lateral Proximal-Distal ACL limits A/P - KT2000 surgical tool –Rotation Varus-Valgus MCL/LCL TorsionalMCL Flexion-ExtensionACL

Collagenous Tissue Testing: Summary of Key Points Tissue testing considerations include –Various possible configurationsconfigurations –The method of tissue clampingtissue clamping –Methods to measure stress and strainmeasure –Defining the stress-free statestress-free state –Anelastic and viscoelastic tissue properties, the need to precondition, and the effects of strain rateAnelastic viscoelastic strain rate –The effects of age, injury, immobilization, surgical repair and replacement.ageinjuryimmobilizationsurgical repair replacement