This is an assessor-blinded, cross-sectional study. The procedures conformed to the Helsinki Declaration and were approved by The University of Sydney Human Research Ethics Committee (2016/748). The study protocol was pre-registered on the Open Science Framework (https://osf.io/fejpf/) and written consent was obtained from all subjects.
Thirty healthy subjects were recruited from The University of Sydney through poster advertisements and convenience sampling. Subjects were included if they were at least 18 years old and did not have previous injury or surgery to the right hip or knee. Subjects were screened to ensure there were no other factors (e.g. delayed onset muscle soreness) that prevented the straight leg raise test from being applied. The right legs of all subjects were tested. A sample size of 28 provided 80% power to detect a 5° mean change in hip angle (standard deviation 9°, alpha 0.05) [6].
Protocol
All testing was conducted during a single visit to a university laboratory. Subjects lay supine on a testing bench with their hips in a neutral position and the tested knee in ~ 5° flexion. The back of the ankle was placed on a load cell (XTRAN S1W 250 N; Applied Measurement, Melbourne, Australia) used to measure knee flexion MVC force (Fig. 1). The skin was cleaned and abraded with alcohol wipes before surface electrodes were applied. Two circular surface electrodes (diameter 10 mm, inter-electrode distance 30 mm) were applied to the back of the thigh over the muscle belly of biceps femoris. Two larger rectangular surface electrodes (50 mm by 90 mm, inter-electrode distance 60 mm) were also applied to the back of the thigh, proximal and distal to the circular electrodes. The hamstring muscles were stimulated through the large rectangular electrodes, while biceps femoris electromyography (EMG) signals were recorded through the small electrodes during knee flexion MVCs and during the trial without stimulation. A ground electrode was placed over the patella.
EMG signals were amplified and filtered (gain 200, 30–500 Hz band-pass; GRASS Model 15LT, Rhode Island, USA) before being sampled, along with the force signal, at 2000 Hz using custom-written LabView software (Labview 2013, National Instruments, Texas, USA) and a National Instruments data acquisition card (NI USB-6251, National Instruments, Texas, USA). A nine degree-of-freedom magnetometer (SparkFun SEN-10724, Colorado, USA) was attached to the distal tibia to measure hip joint angle.
Subjects first performed two isometric knee flexion MVCs against the load cell with the knee in ~ 5° flexion. Subjects were instructed to use only their hamstring muscles (not the gluteal or quadriceps muscles) to produce knee flexion maximal contractions. Verbal encouragement was provided during maximal contractions and a one-minute rest was provided between contractions. For each subject, peak forces during the MVCs were used to determine the intensities for continuous electrical stimulation required to achieve 2.5, 5, 7.5, and 10% of knee flexion MVC force. A constant current stimulator was used to deliver the continuous electrical stimulation (50 Hz, Model DS7AH, Digitimer, Hertfordshire, UK). The intensity required to achieve each target force was determined in separate trials, with a one-minute rest between trials. An additional trial without electrical stimulation (i.e. 0% MVC) was included. During this trial, the biceps femoris EMG signal was monitored to ensure subjects were as relaxed as possible.
The five stimulation intensities were applied to each subject in random order. Subjects were instructed to remain relaxed during all trials, and a one-minute rest was provided between trials. At the beginning of each trial, an investigator (LC or JD) started the electrical stimulation (or pretended to do so for the 0% MVC trial), then the blinded assessor (YF) performed a passive straight leg raise test. The right leg was raised while keeping the knee extended, the test was stopped when the knee started to flex, at which point hip range of motion was recorded (Fig. 1).
Hamstring muscle EMG signals from the knee flexion MVC and 0% MVC trials were digitally filtered (30–450 Hz, band-pass) and the root-mean-square EMG over a 50 ms window was calculated. EMG signals recorded during the 0% MVC trial were normalised to EMG at peak force during the MVC trial. Thus, involuntary hamstring muscle activity is reported as a percentage of maximal knee flexion activation.
Statistical analysis
For each subject, linear regression was used to determine the effect of increasing knee flexion muscle activity (at 0, 2.5, 5, 7.5, and 10% of knee flexion MVC) on hip range of motion. Regression coefficients from all thirty subjects were then pooled to determine the overall mean effect (95% CI) of knee flexion muscle activity on hip range of motion. Secondary univariate sensitivity analyses were performed to determine whether age, sex or body mass index (BMI) had effects on hip range of motion. Data were processed and analysed using Python (version 3.5).