Study 3: In-Vivo Experiment Using Multiple Layers Tracking

This study examined local tendon strain across the anterior, middle, posterior at the proximal and the distal aspect of the patella tendon during ramped isometric contractions. The chosen automatic tracking algorithm (i.e. NCC) was used to track multiple regions of interests (ROI) simultaneously arranged in layers with the same block sizes and positions at each tendon layers. Unlike previous experiments, this study compares the results of each layer and the proximal and distal aspect of the patella tendon to initiate a much more detailed understanding of the tendon in injury, repair and also in response to various training interventions.

3.5.1 Participants

Sixteen healthy limbs were used for data collection in this study from healthy male subjects with an average age of 28.0±6.3 years; height of 1.7±0.04m and body mass 79±5.4 kg.

3.5.2 Tracking Regions

The whole length of the patella tendon was imaged (see Figure 3.17) and the

thickness of the tendon was measured at 8–9mm (a). The regions being examined

were at the proximal (b) and distal ends (c). The regional layers for each tendon end

are divided into anterior, mid and posterior (d-f). The peninsula bone is located at the

proximal ends (g) and tibia at the distal ends (h). The line at the top of the image is

the skin layer (i).

3.5.3 Hardware and Software Setup

Unlike the previous study, an upgraded ultrasound system was used for this experiment (MyLab70, http://www.esaote.com/) with a 7.5 MHz 100mm linear array, B-mode ultrasound probe with a depth range of 67mm was used to image the patella tendon in the sagittal plane. The wider ultrasound probe was capable of capturing more width of the tendon compared to the earlier ultrasound probe (see Figure 3.18. The same setup as before was used for assessing the patella region. The images were then captured at 25 frames per second (fps) in DV format, with image size of 800x600 pixels, and stored locally into the storage memory. The captured frames were then transferred to the computer system for the tracking process. Again, as in the earlier experiments, scaling in pixels per mm was determined from ImageJ software by using the known depth of field in the ultrasound images, (1 mm = 11 pixels in the x and y directions) and utilised as a calibration factor in the automated tracking system to ensure equivalent pixel to mm ratios.

Figure 3.18: Comparisons between a) 40mm probe transducer and b) 100 mm probe transducer. Image produced by the latter probe has a higher depth and broader viewing range. 40mm 100mm a)   b)   Probe Image

The transducer probe was fixed statically at the skin surface, similarly to the previous study. Also, the torque output during isometric quadriceps contraction was determined using an isokinetic dynamometer with the participant in a seated position. The same knee setup was set at 90° flexion and hip at 85°, and a lever attachment cuff was also placed on the lower leg at ~3cm just above the medial malleolus. The tendon was imaged during ramped voluntary isometric contractions (3-4 seconds). The maximal isometric quadriceps contraction efforts were repeated three times to ensure tendon preconditioning prior to the test. Participants performed ramped isometric contractions from 0% MVC (rest) to the maximum (100% MVC) over a 3s to 4s period. Three trials were repeated with 180s rest between contractions. The mean values of strain for the three contractions were used for subsequent analysis.

The EMG of the long head of the biceps femoris (BF) muscle was evaluated in order to determine the level of antagonistic muscle co-contraction during the isometric knee extension (S. Pearson et al., 2006). The assumptions were that BF was representative of its constituent muscle group (Carolan et al., 1992) and that the biceps femoris EMG relationship with knee flexor torque was linear (Lippold, 1952). Three maximal isometric knee flexion contractions were carried out obtaining the EMG at maximal flexion torque. The root mean square EMG activity of the biceps femoris during knee extension was divided by the maximal flexor EMG. Then the maximal flexion torque was multiplied by this value to determine co-contraction torque. The patella tendon force was finally determined by dividing the total torque by the patella lever arm as determined from the literature (Krevolin et al., 2004, Tsaopoulos et al., 2006).

Figure 3.19 shows the regional tracking of ROIs where R1 and R2 are the

arbitrary pixel regions in the tendon arranged into layers; anterior (a), mid (b) and

posterior (c). The ROIs are marked on a typical tendon excursion on both proximal

(A), and distal (B) ends showing shift in ROI's from the resting tendon, at 50% and

100% MVC. The vertical dotted lines show the initial positions of the ROIs. The arrows show the movement of each ROI during the tracking experiment.

Figure 3.19: Regional tracking of the patella tendon.

The resultant displacement for each layer is measured as follows: Δd= (xR2−xR1) 2+ (yR2−yR1) 2

(

)

(

)

where is the change of length, (xR1,yR1) is the position of first ROI, (xR2,yR2) is

theposition of second ROI, is the initial frame and is the subsequent frame. The

strain is measured as follows:

ε =Δd

Δi (3.8)

where is the strain measurement, is the change of length, is the initial

distance between R1 and R2.All the initial proximal and distal regions were aligned

vertically to enable quantification of any differences in regional strain within a localised site of the tendon.

Δd f1 fn ε Δd Δi a    b    c