Ex vivo Mechanical Loading of Tendon

My name's David Rempel. I'm a faculty in the Department of Bioengineering at u at the University of California at Berkeley. The experiment you're gonna see involves stretching tendons and really the purpose is to try to underline the, understand the underlying mechanism to tendon disorders at the wrist and elbow.

These tendon disorders are common among athletes and workers who do repetitive work with their hands. Krish Sundi, one of the doctoral students in our laboratory, will demonstrate several features of the system, specifically the measurement of the cross-sectional area of the tendon. He'll load the tendons into the actuators.

He'll demonstrate the system loading the tendon cyclically over a 24 hour period and then remove the tendon from the system. Okay, so this is the loading system that we've designed and built. As you can see, it consists of six actuators.

These are high precision actuators, which are capable of applying displacement of one micrometer in series with the actuator is a load cell to record loads applied to tendons. The in series with the load cell is, are these clamps which extend away far enough so that they can be submerged in media in order to keep tendons viable. This entire system would then be placed in a water bath to maintain the media at 37 degrees Celsius.

What I'm gonna show you right now is how I would go about loading a tendon into this system. The first step we need to do is measure the cross-sectional area of the tendon so that we can apply the appropriate stresses. Can you pass me the tube with the tendon?

Now, these tendons have been harvested from a New Zealand white rabbit. They have, they're just been brought over, so they're still fresh within less than two hours of have the rabbit being euthanized. A portion of the tendon whose cross-sectional area that I'm measuring is along the thinnest part of the tendon.

This is because this is the point where the greatest stresses will be, will be resulting because of the low, smallest cross-sectional area. Can you now put the weight on top of the piston? This now we wait 30 seconds in order to read the, the thickness read 1.3.

Can you lift the piston and replace the tendon back in the media? And I'm going to unmount dismount the actuator in which I'm going to load the tendon. So now I'm going to mount a tendon into the clamps.

Here we can have a closeup of the actuator. You can see the, the actuator itself, the load cell, and the clamps, which extend away These pads cause pads. I would slice approximately half an inch to three quarters inch wide and approximately one and a half inches long.

Then take these gauze pads and dip them in the media in order to wet them. It makes the rolling process easier And place these rolled gauze strips In the grooves of these serpentine clamps. I'm now going to remove the tendon from the media.

This end is the insertion site of the tendon of the flexor tendon into the end of the, the, the digit of the rabbit. This region here is a fiber cartilage region. The tendon is placed in such a position such that the fiber cartilage region is clamped within the proximal clamp.

The tenant is then clamped, and again, the screws are tied into one newton meter. Then the excess tendon is cut, so now the actuator is secured into the brace and the entire tendon and clamps are submerged. In modified media, The same procedure would be used to mount tendons on subsequent actuators.

As you can see, we have a total of six, so we can do this with six individual tendons. This entire system is then lifted and placed into a water bath in order to maintain the media at 37 degrees. So now we're gonna apply a cyclic load to the tendon between 0.5 and five megapascals.

The software we use is lab view and it's custom written. The first thing we do is to precondition the tendon. We apply 20 cycles of cyclic load between 0.5 megapascals and two megapascals.

As for the thickness measured, use it using the four supplied micrometer to put that in. It will cal automatically calculate the cross-sectional area. The program will then instruct the actuator to increase the displacement until a two megapascal load is applied to the tendon.

Here what we're following is the purple trace. Once a two megapascal load is applied to the tendon, the program will then apply a 0.5 megapascal load to the to the tendon. And once these two loads are found, the program will sit cycle between these two loads, updating the position continuously between these two desired loads.

This loading is being done at 10 megapascals per second at a frequency of 0.5 hertz. Once we have preconditioned the tendon, we then seek a 0.5 newton load. We define the length of the tendon under a 0.5 Newton load as the gauge length of the tendon.

Once the program has found this load, we read the position of the actuator and this position, which reads currently six eight hundred and twenty five, is the gauge length of tendon. After we've recorded the gauge length of the tendon, we then return to the cyclic loading program, input our desired values of five megapascals and 0.5 megapascals, input the total number of cycles we wanna load it to, and again, run through the same program in order to, in order to cyclically load the tendon between 0.5 and five megapascals. Again, this loading is done at 10 megapascals per second with a frequency of 0.5 hertz.

So after being cyclically loaded, typically for 24 hours, the actuator is then removed from the system and from here the tendon can be cut and processed as desired. Well, you've just seen our tendons that were harvested sterly from a rabbits. The process you've saw seen was a semi sterile process, but it led to a cyclical loading of the tendons over a 24 hour period.

The tendons are subsequently analyzed using R-T-P-C-R for gene expression. We're also analyzing the tendons for other changes, structural changes, other histologic markers. Ultimately, the purpose of this experiment is determine what effect different loading patterns have on basically the underlying mechanisms of injury to the tendons that might be associated with tendinopathies and tendonitis.