Static loading of the knee joint results in modified single leg landing biomechanics.

BACKGROUND: External loading of the ligamentous tissues induces mechanical creep, which modifies neuromuscular response to perturbations. It is not well understood how ligamentous creep affects athletic performance and contributes to modifications of knee biomechanics during functional tasks. HYPOTHESIS/
PURPOSE: The purpose of this study was to examine the mechanical and neuromuscular responses to single leg drop landing perturbations before and after passive loading of the knee joint.
METHODS: Descriptive laboratory study. Male (n = 7) and female (n = 14) participants' (21.3 ± 2.1 yrs., 1.69 ± 0.09 m, 69.3 ± 13.0 kg) right hip, knee, and ankle kinematics were assessed during drop landings performed from a 30 cm height onto a force platform before and after a 10 min creep protocol. Electromyography (EMG) signals were recorded from rectus femoris (RF), vastus lateralis (VL), vastus medialis (VM), semimembranosus (SM), and biceps femoris (BF) muscles. The creep protocol involved fixing the knee joint at 35° during static loading with perpendicular loads of either 200 N (males) or 150 N (females). Maximum, minimum, range of motion (ROM), and angular velocities were assessed for the hip, knee, and ankle joints, while normalized EMG (NEMG), vertical ground reaction forces (VGRF), and rate of force development (RFD) were assessed at landing using ANOVAs. Alpha was set at 0.05.
RESULTS: Maximum hip flexion velocity decreased (p < 0.01). Minimum knee flexion velocity increased (p < 0.02). Minimum knee ad/abduction velocity decreased (p < 0.001). Ankle ROM decreased (p < 0.001). aVGRF decreased (p < 0.02). RFD had a non-significant trend (p = 0.076). NAEMG was significant between muscle groups (p < 0.02).
CONCLUSION: Distinct changes in velocity parameters are attributed to the altered mechanical behavior of the knee joint tissues and may contribute to changes in the loading of the leg during landing.

Introduction

Knee joint injuries greatly affect athletic and recreational sport populations. Sex-related and sports based factors are the leading determinants of knee injuries [1,2,3,4]. The loading of the knee joint in dynamic sporting activities influences the stresses and strains, which the tissues within and surrounding the knee joint capsule tolerate. The ligaments of the knee joint provide structural integrity to the joint during both passive and active movements [5]. The incidence of injuries to the passive viscoelastic tissues, such as the anterior cruciate ligament (ACL), posterior cruciate ligament (PCL), medial collateral ligament (MCL), lateral collateral ligament (LCL), and menisci are well documented in the literature [6,4,7] as well as the predictive factors leading to injury [8,9].

The injury mechanisms at the knee joint are multifactorial and are complicated due to the requirements of specific movement activities performed in dynamic environments (i.e., athletics venues). Both contact and non-contact activities play critical roles in determining how the knee joint responds to the given loading conditions. The type of training, task, fatigue level of the individual, and anatomic structure all contribute to the potential injury of the knee joint [10,11]. In particular, when landing from a given height, the knee biomechanics are modified to absorb energy to reduce the impact of the contact forces upon the lower extremities [12]. Females are reported to have greater dynamic knee valgus–a potential sign of knee injury at landing–compared to males, and thus a greater potential for knee injury [13,14,15]. Training individuals to land without excess knee valgus has been documented and may contribute to reduced knee joint injuries [16,17]. Nilstad et al.[18] used static laxity of the ligamentous tissues as a predictor variable for knee valgus, but could not conclude this was a factor responsible for increasing likelihood of knee injuries. Others, however, have determined the laxity of the knee joint ligaments contribute to modifications of the neuromuscular control of the knee joint [19,20,21]. The musculotendinous units contribute much more to the stability of the knee joint during dynamic tasks as the forces generated via the muscle are actively engaged in movement of the joint, as well as resistance to external forces acting upon the knee [22].

Neuromuscular fatigue of the muscles surrounding the knee joint and other lower extremity muscles is a contributing factor to knee injury. Lower extremity kinematics and myoelectric activity, collected with electromyography (EMG), significantly change at landing from a jump or drop after fatigue has been induced [23]. When the muscles become fatigued the ability to generate force diminishes, and the internal moments have reduced capacity to resist the external moments applied [24]. The contribution of neuromuscular fatigue to knee joint injury is significant [11], however, it is unclear how the laxity of the ligamentous tissues contributes to the inability of the joint to maintain its integrity [18]. Neuromuscular fatigue requires the passive ligamentous tissues to be strained further to compensate for the deficiencies of the neuromuscular control. These ligamentous tissues are further loaded during the dynamic activities as tension-relaxation or mechanical creep are induced in parallel with neuromuscular fatigue [25]. Further assessment of the contribution of the passive ligamentous tissues during loading tasks is necessary to understand the mechanisms of injury.

Reductions in both force generating capacity and myoelectric activity of the musculature about the knee joint have been documented during isometric knee actions following passive loading of the knee joint capsule [20,21,26]. These tests serve to isolate the knee joint and provide a foundational understanding of the influence of loading schemes to the response of the neuromuscular and musculoskeletal systems. Functionally, little is known how passive loading of the knee joint capsular tissues affects the movement of the joint during dynamic (athletic) activities.

The purpose of this study was to examine the mechanical and neuromuscular responses of the lower extremities to landing perturbations before and after passive loading of the knee joint capsule. It was hypothesized that passive static loading of the knee joint capsule would elicit a reduced myoelectric amplitude response from the surrounding musculature at landing. Further, it was also hypothesized that joint kinematics of the landing leg would compensate for the passive loading applied at the knee joint capsule during landing through changes in kinematics parameters.

Methods

Participants

University students enrolled in kinesiology classes volunteered to participate in this study. Male (n = 7) and female (n = 14) participants (21.3 ± 2.1 years old, 1.69 ± 0.09m in height, and had a mass of 69.3 ± 13.0 kg) were required to be healthy individuals with no medical conditions which would prevent physical activity, involved in regular physical activity (recreational activities at least 3 times/week), not have any trunk or lower extremity disorders, not have an injury to the head, trunk, and lower extremities within the previous 12 months of participation, and if female not be pregnant. The study was approved by the Southern Illinois University Carbondale Human Subjects Committee (#15277). Participants were provided an informed consent document and written voluntary consent was provided by each participant. Additional verbal instructions were provided during the study. Participants were informed that they could withdraw without penalty at any time during the study.

Instrumentation

A 6-camera motion capture system (Qualisys, Gotenborg, Sweden) with Oqus 100 camera sampling at 120 Hz was used to collect movement data. Palpation was used to place individual reflective marker spheres of 14 mm diameter bilaterally over the acromion processes, posterior superior iliac spines (PSIS), and anterior superior iliac spines (ASIS). Unilateral markers were positioned over the sacrum at the S1 Table, and the right leg at the greater trochanter, lateral femoral epicondyle, medial femoral epicondyle, lateral malleolus, medial malleolus, calcaneus, 1st and 5th metatarsophalangeal joints. Two four-marker clusters were position on the right leg at the midline of the lateral thigh and the proximal third of the lower leg and secured with Coband © wrapping tape.

Surface electromyography (EMG) (Motion Lab System, Baton Rouge, LA, USA) was used to collect muscle activity from the right thigh musculature surrounding the knee. The skin was abraded and then cleaned with isopropyl alcohol. The 0.02 m diameter stainless steel electrodes have a fixed center to center distance of 0.02 m and bipolar configuration, and were positioned distal to the motor point of each muscle group and aligned parallel with the muscle fibers. Myoelectric signals were collected from the muscles rectus femoris (0.10 m distal from the right ASIS), vastus lateralis (0.10 m proximal and lateral form the patella), vastus medialis (0.10 m proximal and medial to the patella), semimembranosis (~0.20 m proximal to the medial femoral epicondyle), and the biceps femoris (~0.20 m proximal to the lateral femoral epicondyle) [27,26]. Surface EMG signals were bandpass filtered at 20–500 Hz with a common mode rejection ratio of > 100 dB at 60 Hz, an input impedance of > 100MΩ, and collected at 1200 Hz.

Kinetic data were collected with a 6 degree of freedom force platform (OR-6, AMTI, Watertown, MA, USA) with dimension 0.45 m x 0.5 m embedded and flush with the laboratory floor. Force data were collected at 1200 Hz. Kinematic, EMG, and kinetic data were collected using the Qualisys Tracking Manager (QTM) software interfaced with a USB 2533 12 bit A/D board (Measurement Computing, Inc., Norton, MA, USA) and save for future processing.

Protocol

Participants warmed up by walking on a motorized treadmill at their self-selected speed for 10 min. Kinematic markers and EMG electrodes were placed upon the participants after the warm-up. Participants performed single leg drop landings from a height of ~ 0.30 m using the right leg (Fig 1). Leg dominance was determined by asking the participants which leg they would use to kick a ball. All participants indicated right leg dominance. Participants began by standing on two legs on top of a box situated 0.10 m horizontal from the force platform. They were instructed to lean forward leading with the right leg in order to drop onto the force platform. Once they landed, the participants were instructed to maintain their one-legged stance and stand erect for 5 s. The hands were positioned on the iliac crests to control arm movements. Participants were given up to 10 practice trials to acclimate to the drop landing task. Participants performed up to 10 trials of drop landings before and after the knee joint capsule was loaded. At least 1 min of rest was provided between trials to reduce the influence of fatigue. Landing trials where the participants either jumped, stepped down, or could not maintain balance at landing were discarded and additional trials were performed. Of the10 trials recorded, the middle 5 trials were used for analysis.

Fig 1

Drop landing sequence.

An example of the sequence in the drop landing protocol.

After the initial drop landing trials, participants were positioned into a high-back chair of a Biodex system 3 dynamometer (Shirley, NY, USA). Participants were positioned with their trunk in an upright erect position with the hips in 90° flexion. Straps were placed across the chest and proximal thigh to reduce movement during the exercise. Then an attachment arm was secured to the dynamometer axis, which was aligned with the lateral femoral epicondyle of the right leg. The attachment arm was secured to the leg 0.05 m proximal to the lateral malleolus. Ramped maximal voluntary isometric efforts (MVIE) of 5 s were performed 3 times each with the knee at 90° flexion for extension trials and 45° flexion for knee flexion trials (full knee extension is 0°), respectively, with a 60 s rest between efforts. A 10 min rest period was performed after the last MVIE. The leg was then positioned so that the knee was flexed to 35° with reference to the anatomical position [20]. A cuff was secured 0.03 m distal to the femoral epicondyle and surrounded the proximal leg. A pulley system was configured to allow a cable to fit perpendicular to the leg and around the cuff. The cable was used to pull the leg anterior relative to the femur with a load of either 200 N (men) or 150 N (women) (Fig 2). This protocol has been reported to increase laxity of the tissues surrounding the knee joint capsule and potentially the ligaments within the capsule [20]. The knee joint was statically loaded for 10 min. Surface EMG was used to ensure a low level of muscle activity relative to the MVIE was maintained during knee loading. Immediately after the loading protocol was completed, participants performed additional drop landing trials.

Fig 2

Set up for the static loading protocol.

Depiction of the static loading of the knee for each participant. The striped box represents the load.

Data processing

The EMG signals collected during the static loading were centered, full wave rectified, and low pass filtered at 3 Hz with a fourth order zero-lag Butterworth filter. The EMG signals collected during drop landing trials were centered, rectified, and then low pass filtered at 5 Hz with a fourth order zero-lag Butterworth filter. All EMG data were then normalized to the maximum EMG value attainted during MVIE.

Force data were processed with a low pass Butterworth filter set at 60 Hz using the MotionMonitor System software (MotionMonitor, Chicago, IL, USA). A vertical ground reaction force (VGRF) threshold of 20 N was establish to determine the onset of load acceptance at landing. Force data were reduced to 120 Hz to coincide with the kinematics data.

Kinematics data were processed using the MotionMonitor software (MontionMonitor, Chicago, IL, USA). Kinematics data were smoothed with a fourth order low pass zero-lag Butterworth filter set at 10 Hz. A static reference file was used to determine lower extremity segment and joint angles using the right-hand configuration (x-anteroposterior, y-mediolateral, z-vertical axes, respectively). Based upon the static reference file, segmental coordinate systems were established and used in determining relative angles of adjacent segments. Further, Euler angle calculations were performed to determine segment orientations, which contributed to joint angular rotations (X–frontal plane, Y–sagittal plane, and Z–transverse plane) of the distal segments relative to the proximal segments.

Data analysis

Kinematics data were evaluated in all three planes of movement during the landing phase of the drop landing. Landing phase during the drop landing was assessed as the onset of the force platform threshold until the maximum knee flexion angle was attained. Joint angle parameters (maximum, minimum, and range of motion) were assessed using the right-hand rule as follows: at the knee joint, flexion-extension (about the y-axis), internal-external rotations (about the z-axis), and mediolateral rotations (about the x-axis). At the hip joint, hip flexion-extension (about the y-axis) and hip adduction/abduction (about the x-axis) were evaluated. Ankle plantarflexion and dorsiflexion movements were assessed as these movements have been reported to align with maximum knee joint flexion at landing [28]. Angular velocities (maximum and minimum) of the three knee joint rotations were also calculated to determine differences between single leg landing trials before and after static knee loading.

Kinetic variables of concern were the average and maximum VGRF, force profile of the first 200 ms of landing, and the rate of force development (RFD). Average VGRFs were compartmentalized into four 50ms time intervals (0–50, 51–100, 101–150, and 151–200 ms) to assess the pattern of forces at landing. The RFD was calculated as the difference in maximum VGRF and the VGRF at landing divided by the time between the maximum VGRF and VGRF at landing.

Surface NEMG recordings during the static knee loading protocol were averaged the first 30 s of each minute of the 10 min loading period. This was performed to ensure minimal muscle activity during the loading. Additionally, processed NEMG signals were assessed during the first 200 ms of the landing phase, as well as 200 and 100 ms prior to initiation of the landing to compare landing before (pre-loading landing condition) and after (post-loading landing condition) static knee loading. Pre-and post-loading landing condition NEMG signals were used to determine the preparation of the muscles surrounding the knee joint to the landing (feed-forward control at 200 and 100 ms). The 200 ms after landing was compartmentalized into four 50 ms intervals (0–50, 51–100, 101–150, and 151–200 ms) which were used to average the NEMG signals for evaluating the trend of the neuromuscular activity at landing.

Statistical analyses

All statistical testing was performed with SPSS v 22.0 (Chicago, IL, USA). Angular displacement and velocity variables (minimum, maximum, and range of motion) were analyzed using a one-way (condition) analysis of variance (ANOVA). A one-way ANOVA was performed to assess average muscle activity at each minute of static loading. Average and maximal NEMG values during landing were analyzed with a 2 factor (condition x muscle) ANOVA. A three-factor, muscle x condition x time interval (5 x 2 x 6) ANOVA was performed on the average NEMG, which included analyzing the pre-landing times at 200 and 100 ms. Average and maximal forces and RFD data were each analyzed with a one-way ANOVA (condition). A 2-way ANOVA (condition x time interval) was used to assess average VGRF data during the landing phase, while one-way ANOVAs were used to compare maximal VGRF values between conditions. Tukey post-hoc comparisons were performed when significant effects were present. A Mauchly’s test of Sphericity was performed to assess the normality of the data. A Greenhouse–Geisser test was applied when normality was not attained. The level of significance was set at p ≤ 0.05.

Results

Electromyography

Average NEMG values did not significantly change during the 10 min of static loading. The average activity for each minute was under 5% of the MVIE, indicating minimum active neuromuscular response to the external load (Table 1).

Table 1

Average NEMG from static loading.

	Muscle Group (% MVIC)
Time (min)	RF	VL	VM	SM	BF
1	3.72 (3.8)	4.15 (2.6)	3.59 (4.1)	4.87 (4.3)	2.80 (2.0)
2	3.67 (4.2)	3.79 (2.0)	2.67 (1.4)	4.46 (4.0)	3.45 (2.7)
3	3.57 (4.3)	3.92 (2.5)	2.78 (1.5)	4.06 (3.4)	3.09 (2.5)
4	2.99 (3.2)	4.08 (2.4)	2.73 (1.5)	3.52 (2.5)	2.82 (2.3)
5	3.00 (3.2)	4.03 (2.7)	2.73 (1.5)	3.71 (2.9)	2.71 (2.4)
6	2.95 (3.1)	3.74 (2.1)	2.66 (1.4)	3.49 (2.5)	2.56 (2.2)
7	2.79 (2.9)	4.11 (2.9)	2.60 (1.4)	3.53 (2.5)	2.79 (2.3)
8	2.76 (2.9)	3.90 (2.1)	2.89 (1.5)	3.49 (2.5)	2.57 (2.1)
9	2.75 (2.9)	4.25 (2.8)	2.71 (1.5)	3.43 (2.4)	2.50 (2.0)
10	2.74 (3.0)	3.84 (1.8)	3.06 (2.3)	3.32 (2.4)	2.34 (1.8)

Mean (± sd) of normalized surface electromyography as a percentage of MVIC from the three quadriceps and two hamstring muscles during static loading of the knee joint capsule. RF = rectus femoris; VL = vastus lateralis; VM = vastus medialis; SM = semimembranosus; BF = biceps femoris.

Overall average and maximum NEMG during landing

Average NEMG values did not change between conditions (p > 0.55), but were significant between muscles (F4,199 = 5.347, p < 0.01). There was no significant condition x muscle interaction effect (p > 0.87) (Table 2). Maximal NEMG values were not significant between conditions (p > 0.34), but were significant between muscles (F4,180 = 9.553, p < 0.01) (Table 2). There were no significant interaction effects present (p > 0.97).

Table 2

NEMG from thigh muscles during pre- and post-loading landing conditions.

	Maximum NEMG		Average NEMG
Muscle	Pre	Post	Pre	Post
RF*	1.15 (0.62)	1.18 (0.55)	0.451 (0.30)	0.477 (0.28)
VM^	0.43 (0.45)	0.45 (0.55)	0.284 (0.32)	0.241 (0.23)
VL†	0.98 (0.62)	1.04 (0.50)	0.506 (0.37)	0.619 (0.55)
BF	0.87 (0.56)	0.97 (0.62)	0.385 (0.28)	0.391 (0.28)
SM‡	0.64 (0.31)	0.809 (0.56)	0.307 (0.15)	0.342 (0.29)

Mean (sd) maximal and average NEMG as a percentage of MVIC from the three quadriceps and two hamstring muscles during pre- and post-loading landing conditions.

*indicates RF significantly greater than VM (p < 0.01)

†indicates VL significantly greater than VM, BF, and SM muscle groups (all p < 0.02)

^indicates VM significantly less than all other muscle groups (all p < 0.03)

‡Indicates SM significantly different than RF, VL, and VM (all p < 0.03)

A significant time interval x muscle interaction was present (F20, 1189 = 1.951, p < 0.01) when observing the average NEMG signals at landing. Post-hoc analysis indicated a significant difference between time intervals (p < 0.001) and muscle groups (p < 0.001). There were no significant condition effects (p > 0.27), nor condition x time interval (p > 0.99) or condition x muscle interaction (p > 0.99) effects present (Fig 3).

Fig 3

Average NEMG at 200ms prior to and at landing.

Mean (sd) average NEMG from pre and post-loading landing conditions for rectus femoris (RF), vastus lateralis (VL), vastus medialis (VM), biceps femoris (BF), and semimembranous (SM) muscle groups. NEMG activity is provided in 50ms intervals during the first 200ms of landing. NEMG values are also provided 200ms and 100 ms prior to the landing.

Kinematics

Maximum and minimum angular displacement data from the hip, knee, and ankle joints are provided in Table 3. Although not significant, a trend was present between conditions for ankle flexion maximum (p < 0.071). All other maximum and minimum angular displacement data were not significant between conditions.

Table 3

Joint angular displacements at landing.

	Maximum		p-value	Minimum		p-value
	Pre	Post		Pre	Post
Hip flexion	22.0 (6.8)	19.7 (9.2)	0.38	9.2 (3.0)	6.2 (6.6)	0.15
Knee flexion	60.8 (7.8)	60.9 (8.3)	0.97	16.4 (4.8)	15.7 (9.7)	0.67
Ankle flexion	1.9 (16.9)	-1.9 (12.6)	0.071	-52.7 (14.5)	-51.4 (15.4)	0.61
Hip Abduction	14.0 (8.9)	11.3 (8.6)	0.13	-7.6 (13.9)	-6.4 (11.0)	0.59
Knee abduction	-4.0 (8.3)	-4.2 (6.2)	0.94	-30.5 (11.9)	-32.7 (14.1)	0.29
Knee rotation	16.2 (5.8)	16.7 (9.5)	0.77	1.7 (6.9)	2.8 (2.4)	0.48

Mean (sd) maximum and minimum angular displacements (°) at the hip, knee, and ankle joints during pre- and post-loading landing conditions. Ankle dorsiflexion is represented as positive, and plantarflexion is noted as negative.

Table 4 provides angular velocity data measured from the hip, knee, and ankle joints. Hip abduction maximum velocity was significant between conditions (F1,3 = 43.5, p < 0.007). A significant difference between conditions was present (F1,9 = 7.963, p < 0.02) for minimum knee flexion velocity. Minimum knee abduction velocity was significant between conditions (F1,9 = 19.35, p < 0.002). All other angular velocities in sagittal, frontal, and transverse planes were not statistically different between conditions.

Table 4

Joint angular velocities.

	Maximum		p-value	Minimum		p-value
	Pre	Post		Pre	Post
Hip flexion	204.9 (76.4)	227.0 (46.5)	0.43	-60.9 (62.2)	-65.5 (73.5)	0.78
Knee flexion	527.9 (131.6)	544.1 (168.7)	0.59	-14.9 (25.2)	-27.9 (34.2)	< 0.02
Ankle flexion	792.3 (104.9)	544.1 (168.7)	0.66	-4.7 (21.3)	-17.0 (41.5)	0.21
Hip Abduction	87.1 (56.9)	56.9 (63.4)	< 0.007	-304.2 (101.2)	-291.7 (62.6)	0.57
Knee abduction	90.0 (48.4)	99.7 (50.5)	0.53	-528.5 (127.5)†	-399.9 (129.3)	< 0.002
Knee rotation	306.3 (99.9)	276.4 (89.5)	0.30	-125.9 (51.4)	-141.5 (45.2)	0.46

Mean (sd) maximum and minimum angular velocities (°/s) at the hip, knee, and ankle joints during pre- and post-loading landing conditions. p < 0.05 is indicated in bold.

Table 5 provides ROM data for the hip, knee, and ankle joints. Ankle ROM was significant between conditions (F1,166 = 7.904, p < 0.006). No other ROM variables were significantly different between conditions.

Table 5

Joint ranges of motion.

Angle (°)	Pre	Post	p-value
Hip flexion	15.9 (8.6)	16.6 (8.8)	0.64
Knee flexion	46.3 (9.8)	46.8 (9.8)	0.73
Ankle flexion	56.2 (8.5)	52.6 (8.5)	< 0.006
Hip abduction	19.3 (11.1)	18.5 (11.4)	0.71
Knee abduction	21.5 (13.0)	21.9 (12.8)	0.82
Knee rotation	14.5 (8.1)	14.2 (8.1)	0.79

Mean (sd) values of ROM of the hip, knee, and ankle joints during pre- and post-loading landing conditions. p < 0.05 is indicated in bold.

Kinetics

Maximum VGRFs were not different between pre-loading and post-loading landing conditions (1550.5 ± 84.6 N vs.1548.1 ± 55.8 N, p > 0.91). The average VGRF measures over the first 200ms of contact at landing were significant between pre-loading and post-loading landing conditions (1297.1 ± 392.4 vs. 1231.3± 392.4 N; F1,795 = 5.593, p < 0.018). A significant difference in the maximum VGRFs between time intervals was present (0-50ms: 940.8 ± 322.7; 51-100ms: 1661.4 ± 491.2; 101-150ms: 1355.5 ± 388.7; 151-200ms: 1099.7 ± 350.2 N) (F3,787 = 128.217, p < 0.001) (Fig 4). Post-hoc comparisons indicated the VGRF was significantly different between each time interval (p < 0.001). No significant condition x time interval interaction effect was present (p > 0.56). The rate of force development was not significantly different between conditions, but a trend was present (pre: 16,602.0 ± 1057.0 N/s, post: 17,368.0 ± 1447.6 N/s, p < 0.076).

Fig 4

Interval VGRF.

Mean (sd) maximal vertical ground reaction forces measured at 50 ms intervals from initial landing to 200ms during each condition (pre and post). * indicates significant difference between each 50ms time interval (p < 0.001).

Discussion

The aim of this study was to assess the neuromuscular and kinematics responses of the landing leg during single-leg drop landings before and after passive static loading of the knee joint capsule. Based upon previous research involving passive loading of the knee joint capsule, it was believed that neuromuscular and biomechanical behaviors would be modified in the lower extremity at landing. The reasoning for this study was twofold: 1) mechanical loading of the viscoelastic passive tissues is known to influence mechanical behavior changes of the affected (loaded) tissues, as well as the EMG response of the surrounding muscles, and 2) in the absence of neuromuscular fatigue it is not known how the lower extremity will respond to a perturbation given during a functional activity once passive loading of the knee joint is performed. The initial hypothesis predicted a reduction in EMG amplitude of the muscles surrounding the knee joint at landing after passive knee joint loading. However, this hypothesis was not supported based upon the results. There were no significant neuromuscular changes between pre- and post-loading landing conditions. It was also assumed that the neuromuscular system would compensate for the reduced mechanical tension within the passive connective tissues to increase coactivation in the drop phase prior to landing. This, however, was not substantiated in the data and cannot be considered a control mechanism of the leg at landing in this study. The second hypothesis regarding compensation of joint motion due to the passive loading at the knee joint capsule was partially supported. Distinct modifications to hip, knee, and ankle kinematics during landing resulted from the static loading. Further, the overall average VGRF decreased after static loading, while no change in the maximal force was evident.

Neuromuscular control determined the response of the landing leg during the drop landing. Low level myoelectric activity from both hamstrings and quadriceps muscles groups assisted in preparing the leg for landing. This feed-forward mechanism allows the neuromuscular system to engage immediately once contact with the support surface is initiated to prevent the leg from buckling. Once landing occurred, greater muscle responses were observed to reduce joint angular motions. There were significant differences between the muscles groups at landing. Overall, maximum NEMG values were greatest in the RF group and least in the VM muscles group. However, average NEMG values were greatest from the VL group, and least in the VM group. This is expected as the VL muscles group has been reported to be activated at higher relative levels during similar tasks [22].

The descent phase of landing requires knee extensor muscles to perform eccentric actions to diminish flexion of the knee joint. The NEMG signals of all muscles examined increased as the knee approached maximum flexion. The contribution of the VL and RF muscles were much greater than that of the VM, indicating differential control within the knee extensors. This may indicate the inability of the VM muscle group to provide a primary role in joint stability, which may be due to architectural factors [29]. The increased myoelectric amplitudes are expected from the knee extensors, but not necessarily from the knee flexor muscles. The SM muscle group showed high activation levels in both conditions indicating that this muscle group was more actively involved in the control of leg mechanics compared to the BF muscle. There are two explanations which exemplify the activation of the hamstrings at landing: 1) the hamstring muscle activities are indicative of control at the hip joint to reduce hip joint flexion motion during the landing phase, and 2) sufficient hamstring muscle activity is required to compensate for anterior translation of the tibia [30].

Reduced BF muscle activity is suggested to increase knee internal rotation in a small sample of female athletes [31]. In patients who have undergone ACL reconstruction, modifying landing instruction to increase knee flexion at landing was reported to also reduce BF activity [27]. Although the BF activity was relatively lower than other muscles surrounding the knee, this did not influence overall knee flexion at landing (Table 3).

Compensatory changes in the movement velocities at the knee and hip during the landing phase highlight the modified control the musculoskeletal system uses to respond to the dynamic loading. First, the rate of hip abduction at landing was significantly reduced (Table 4) even though the displacement of the hip joint during landing did not change between landing conditions (Table 3). Reduced hip abduction angular velocity suggests a potential increase in hip adductor contribution during the landing. This negative hip abduction velocity may compensate for the mechanics observed at the knee joint. The ability of the leg to absorb the shock at landing may have been due to greater emphasis of control at the knee as greater negative knee flexion angular velocity was observed. In addition, a reduced knee abduction negative velocity would indicate a greater control of frontal plane knee rotations and less mechanical energy being absorbed. Norcross et al. [32] reported knee landing kinematics differences between ACL injury risk groups and noted greater increased ligament loading with greater energy absorption. It is possible that the mechanical energy absorbed by each joint at landing was modified and could explain how control at each joint was performed.

The knee joint kinematics may have influenced the range of motion observed from the ankle joint after the passive loading. Overall ankle joint range of motion decreased, while no maximum or minimum angular displacement measures were different between the landing conditions. Fong et al. [28] noted that passive ankle joint range of motion was related to greater knee flexion at landing leading to reduced stress in the ACL, specifically. Although greater range of motion can reduce the forces acting upon the joints at landing, Butler et al. [33] suggest increased joint stiffness is important for successful landing mechanics. However, increased limb stiffness is also a factor in potential lower extremity injury, particularly in female athletes [34,35]. Additionally, maximal hip abduction velocity decreased, minimum knee flexion angular velocity increased in magnitude, and knee abduction velocity decreased indicating potential neuromuscular control enhancement of the muscles surrounding the joints.

Significant reductions in the average VGRF after static loading of the knee joint was present. Initially, it is possible that the landing style changed between conditions, however, there was no kinematics evidence to suggest foot position changed at the initial contact with the support surface. Although a reduced range of motion at the ankle joint in plantar-dorsiflexion was observed, this was not believed to influence the landing, especially within the first 10 ms of the landing, which is a critical time period of knee injuries at landing. Kernozek et al. [14] observed a non-significant trend of reduced maximal VGRF, as well as reduced internal joint reaction forces during drop landings performed after fatiguing the thigh musculature. Similarly, Laughlin et al. [36] reported reduced maximal VGRFs and maximal ACL forces when female participants were instructed to land with greater knee flexion during drop landing. They observed kinematics differences at initial contact and maximal ACL force from the hip and knee joints which explained their findings.

Although not tested, the stiffness of the leg influences the ability of the system to resist external loads applied. In particular, the musculotendinous stiffness influences the knee joint loading and ability to dissipate mechanical energy. Greater joint stiffness at landing when the knee is more extended leads to increased injury potential [37,38]. Hamstring musculotendinous stiffness has been reported to reduce the loading of the ACL and limit frontal plane rotations [39]. This is significant as the current study was implemented to reduce stiffness in the tissues within and surrounding the knee joint. In addition, joint stiffness has been reported to be greater in females compared to males at landing [35]. This may serve as an initial protective mechanism for the joint at landing, but may act to increase the chances of knee ligament injury.

Knee joint loading

Isolation of the knee joint utilizing specific loading schemes to assess the neuromuscular responses of the surrounding joint musculature provides biomechanical information of the factors associated with knee joint injury mechanisms, in the absence of neuromuscular fatigue. When muscles become fatigued more of the load/stress is transferred to the passive viscoelastic tissues to maintain joint integrity during functional movements. Although not a functional loading scheme, the passive loading implemented in the current study has been shown to elicit creep behavior of the tissues within and surrounding the knee joint capsule [19,20]. Evidence of the influence of these mechanical creep experiments has provided mixed information, but this is also dependent upon the specific intentions of each study. Cheng et al. [19] initiated posterior loading of the tibia to elicit posterior cruciate ligament creep and reported reduced co-activation of the antagonist thigh muscles during knee extension activities. Chu et al. [20], however, noted increased force and agonist activation during maximal effort knee extension exercises with no changes in antagonist (hamstring) activities. Further evidence of passive tissue loading within and surrounding the knee joint demonstrates reduced agonist and antagonist muscle activities in maximal efforts [21,26], indicating a potential neuromuscular inhibition which may impede function of the muscle during activity. Specifically, Nuccio et al. [21] report significant reductions in the biceps femoris muscle activity after cyclic loading during both knee flexion and knee extension static efforts.

It must be emphasized that isolated loading of specific tissues, such as the ACL or PCL in the knee joint capsule, are not directly linked in in vivo studies. Unlike animal models where tissues can be isolated for perturbation/loading to determine the effects of mechanical manipulation of the specimen [40,41,42], there are factors which constitute how human models can be interpreted. Loading of the knee joint involves applying mechanical creep to the surrounding musculotendonous units, ligamentous tissues, meniscus, and other connective tissues which assist in maintaining the functional dynamics of the knee during physical activity. Particularly when applying these anteriorly directed loads, the musculotendonous units of the hamstrings muscle group can be strained leading to potential modifications in the activation level and stiffness of the muscle [40]. Shear stress of the meniscus during anterior loading is reported to differ between femoral and tibial anterior and posterior attachments, as well as medial and lateral attachments leading to an overall disparity in load distribution in ACL-deficient knees [43].

Limitations

There were limitations to this study which need to be addressed. Relatively moderate loads were applied to the knee joints. This was performed to elicit a creep behavior in the viscoelastic connective tissues, as shown by Chu et al.[20]. Increased loads applied to the knee joints may increase the creep response and this may modify the results presented in this paper to coincide with loads incurred during athletic events. The dominant leg was assessed in the current study. Injuries occur in both dominant and non-dominant leg, and the responses to similar loads may be different between these limbs [44,45] Therefore, additional measures are required to assess both knees in future studies. Further, sex-specific differences between men and women need to be examined to better understand how these loading schemes influence neuromechanical responses.

Conclusions

Implementation of a static load to the knee joint capsule modified movement parameters during a drop landing performance. Tissue-level behavioral changes may be present to influence how the lower extremity joints respond to dynamic loading. Neuromuscular modifications were not present between the landing conditions indicating that this loading scheme does not result in altered neuromuscular control. Additional research is warranted to examine potential modifications to the loading schemes to further understand how the neuromechanics of the lower extremities are modified when controlling for fatigue.

Mean (sd) average EMG at pre- and post-loading landing at 200 and 100ms prior to landing, and at 50 ms intervals at landing.

(DOCX)

Click here for additional data file.

Mean (sd) maximal EMG from each muscle group during pre- and post-loading landing conditions.

(DOCX)

Click here for additional data file.

Mean (sd) maximal VGRFs at each 50ms interval of pre- and post-loading landing up to 200 ms.

(DOCX)

Click here for additional data file.

19 Sep 2019

PONE-D-19-17819

STATIC LOADING OF THE KNEE JOINT RESULTS IN MODIFIED SINGLE LEG LANDING BIOMECHANICS

PLOS ONE

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Reviewer #2: Partly

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Reviewer #1: Yes

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Reviewer #1: Research is conducted according the paper guidelines. Methodology and project design are set correctly. The only objection is the small number of subjects who participated in the study. It is not possible to make a quality conclusion based on such a number. It would be necessary to increase the number and adjust the statistical analysis in order to reach a good conclusion.

Reviewer #2: Review the manuscript titled “Static loading of the knee joint results in modified single leg landing biomechanics”

Manuscript Number: PONE-D-19-17819

The aim of this study was to investigate the muscular, kinetic and kinematic response of the lower limb during landing following anterior passive loading of the capsuloligamentous structures of the knee joint. The experimental setup used in the present study was similar to the one used by Chu et al (2003) who also investigated the effect of creep that developed in the anterior cruciate ligament due to prolonged static loading on the reflexive activation of the associated musculature.

Even though the scientific value of this study is unquestionable, as it enlightens the mechanisms that underlie knee injuries, there are some issues that require author’s respond, particularly with regard to the method, the statistical analysis of the research data as well as the presentation and discussion of the results, before the study is considered for publication.

General comments

Method

• The authors should justify the choice of loads that they implemented for static loading of the knee joint as these could be considered light to mild in comparison to the loads that are developed in sporting activities such as during landing. This issue should be discussed more thoroughly and possibly introduced as one of the limitations of the present study.

• The authors should also justify their choice to use the dominant leg (the one the participants use for kicking a ball) in their study as previous studies have demonstrated significant differences between the dominant and non-dominant leg regarding knee joint stability during unilateral landing (Ludwig et al 2017; Herrington 2011).

• Given the complexity of the experiment, it is suggested that the researchers provide photographic material for the experimental set up and procedure.

Statistical analysis

The general sense that someone gets from reading this manuscript is data over-analysis. The authors performed several statistical tests in order to identify potential effect of static loading of the knee’s passive structures during landing. However, there is a mismatch (in my opinion) between the statistical tests that were performed, the presentation of the Results and the data listed or depicted in Tables and Graphs, respectively. More specifically:

• The authors performed a Two-Way (condition x muscle) ANOVA to compare average and maximum normalized EMG of each muscle involved during landing between pre and post loading conditions (the results are listed in Table 1); they also performed a 5 (muscles) x 2 (conditions) x 4 (time intervals) Two-Way ANOVA for average normalized EMG (the results are graphically presented in Figure 1). It is clear that in the latter case the authors attempted to investigate how muscles’ EMG behave over time during landing; however in Figure 1 the normalized EMG that was recorded before landing (200-0 and 100-0 ms) is also depicted. Why the authors did not include the pre-landing EMG’s in the statistical analysis? Their decision should be is justified and, in any case, they should consider modifying Figure 1, as in its current state it seems that both pre-landing and during landing EMG responses were also compared.

• Similarly, EMG muscles ratios have been compare with two different statistical tests. A muscle ratio x condition Two-Way ANOVA was used to compare average and maximum EMG ratios between the pre- and post-loading condition (the results are listed in Table 3 and 4) and again a 6 (muscle ratios) x 2 (conditions) x 5 (time intervals) Two-Way ANOVA for muscles EMG ratios (results are graphically presented in Figure 2). In this case (i) the authors did not report whether muscles EMG ratios were based on the average or the maximum EMG activity, (ii) they included 5 time intervals whilst the time intervals during landing were 4; which was the 5th time interval? Did the authors include in their analysis a pre-landing time interval (200-0 or 100-0 ms)? And if they do so, why they did not the same for average and maximum EMG activity of the muscles under investigation? (iii) an additional column was added in Figure 2 (the one that represents total EMG ratio for each time interval) but it should be clarified whether or not it was included in the statistical analysis. The authors should consider omitting the “total EMG ratio column” particularly if its scientific value is meaningless as it confuses the reader (see next comment).

• In both aforementioned cases the “overall” average and maximum EMG as well as the “overall” muscle EMG ratios is presented in Tables 2-4. In Figure 2 and Figure 3 a “total” EMG ratio column and a “combined” vertical ground reaction forces column (VGRF), respectively is also presented. The authors should justify whether (i) the “overall”, “total” or “combined” are terms that were used interchangeably, (ii) these terms represent data obtained at the pre-landing and during landing condition (which in my opinion make sense) or data obtained before loading and after loading of the passive knee structures. The legends in Tables 3 and 4 add more confusion to the reader as it is not clear whether the data listed in Tables represent “… average and maximal muscle ratios during landing phase between pre and post landing conditions” or “… loading conditions”. In any case the authors should consider omitting the “overall”, “total” or “combined” data, as in my opinion this information neither adds more scientific value to this research nor it is thoroughly discussed.

• For VGRF the authors (i) reported the test that they used in statistical analysis, (ii) in the Results section they reported no statistical differences between conditions with regard to absolute peak and normalized peak forces but (iii) in the Discussion section they stated that “significant reductions in the average VGRF after static loading of the knee was present” (Page 20, Lines 429-430). This information was probably extracted from the graph in Figure 3 (?), but the authors neglected to report the level of statistical significances either in the text or in the graph.

• The statistical tests used for maximum and minimum angular displacement, angular velocities and ROM comparisons between conditions were not reported.

Discussion

The authors should discuss the limitations of the study and propose future research based on the findings and / or the methodological Inadequacies of the present study.

Minor comments

Page 3, Line 47: The authors should consider replacing “… stress and strains which …” with “… stress and strains, which…”

Page 3, Line 61: The authors should clarify whether they are referring to the anatomical (malalignment of mechanical axes) or dynamic “… knee valgus …”

Page 4, Line 68: The authors should consider replacing “…musculotendonous…” with “…musculotendinous…”

Page 6, Line 126: The authors should describe the technique used, if any, for identification of motor points and/or cite related references.

Page 7, Lines 148-152: The authors should clarify the number of test trials that were actually included in data analysis as according to their protocol 10 test trials were required but eventually “… 5 sufficient trials…” were enough. Does this mean that the average of 5 trials out of the 10 was included in the data analysis and, if yes, which trials were discarded if more than 5 trials were recorded?

Page 13, Line 276: The authors should replace (or not) the “… pre and post landing conditions” with “… pre and post loading conditions” in the legend of Table 3. The same applies for the legend of Table 4 (Page 14).

Page 13, Table 3: The authors should consider omitting the “overall” data (unless they explain and justify its use) since, in my opinion, it is confusing, not related with the initial hypothesis and eventually not discussed, although it appears that is the only data sets that present statistical significances. The same applies for the legend of Table 4 (Page 14).

Page 15, Line 307: The authors should consider replacing “… (p>0.071)” with “… (p<0.071)”

Page 19, Lines 400-401: The authors should consider replacing “… overall knee rotation at landing” with “… overall knee flexion at landing”. They should also consider modifying Table 5 as in this Table the minimum and maximum, but no “overall”, displacement is listed.

Page 19, Line 407: The authors should consider replacing “… (Table 4)” should with “… (Table 5)” as the latter one display data values of joints angular displacements while the former displays data values of EMG muscle ratios.

Page 19, Lines 410-411: The authors should specify the negative values in both the text and in Tables 5 and 6.

Figure 1: The units for NEMG should be added in the vertical axis

Figure 1-3: The author should use the same title in the horizontal axes (Time or Time intervals) in all Figures. The levels of statistical significance may also be indicated in a revised version of the manuscript provided that the proposed amendments have been taken into account.

Figure 3: The legend should be modified according to the other graphs for consistency. For example: Pre-loading � instead of Pre - black

**********

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Reviewer #1: No

Reviewer #2: Yes: Dimitris Mandalidis, Assistant Professor, Department of Physical Education and Sports Science, National and Kapodistrian University of Athens

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14 Nov 2019

Dear Dr. Mirkov,

The suggestions and comments posed by you are the reviewers are greatly appreciated. After reviewing the feedback, the manuscript was revised to better communicate the intensions of the study. A manuscript with the track changes, to show the edits made, and a clean copy of the revised manuscript have been provided. Reponses to the reviewers have been indicated with R, and in bold type.

Thank you.

Response to Reviewers

Journal Requirements:

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Please ensure that your manuscript meets PLOS ONE's style requirements, including those for file naming. The PLOS ONE style templates can be found at

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R: Thank you for this comment. The manuscript has been formatted to the specifications of the journal.

2. Thank you for including your ethics statement: "Southern Illinois University Carbondale Human Subject Committee

Approval # 15277

written consent was provided by each participant"

a. Please amend your current ethics statement to confirm that your named institutional review board or ethics committee specifically approved this study.

b. Once you have amended this/these statement(s) in the Methods section of the manuscript, please add the same text to the “Ethics Statement” field of the submission form (via “Edit Submission”).

For additional information about PLOS ONE ethical requirements for human subjects research, please refer to http://journals.plos.org/plosone/s/submission-guidelines#loc-human-subjects-research

R: The ethical statement has been amended and added as requested (p.5. lines 106-108).

Reviewers' comments:

Reviewer #1: Research is conducted according the paper guidelines. Methodology and project design are set correctly. The only objection is the small number of subjects who participated in the study. It is not possible to make a quality conclusion based on such a number. It would be necessary to increase the number and adjust the statistical analysis in order to reach a good conclusion.

R: Thank you for highlighting this item. It is believed that a quality conclusion can be established, based upon the reported data. The sample size was believed to be sufficient for this study. Areas of analysis have been streamlined to better present the information. As far as the statistical analysis is concerned, the explanation of the statistical procedures have been revised to more accurately describe how the data were assessed (p.11, lines 225-238).

Reviewer #2: Review the manuscript titled “Static loading of the knee joint results in modified single leg landing biomechanics”

Manuscript Number: PONE-D-19-17819

The aim of this study was to investigate the muscular, kinetic and kinematic response of the lower limb during landing following anterior passive loading of the capsuloligamentous structures of the knee joint. The experimental setup used in the present study was similar to the one used by Chu et al (2003) who also investigated the effect of creep that developed in the anterior cruciate ligament due to prolonged static loading on the reflexive activation of the associated musculature.

Even though the scientific value of this study is unquestionable, as it enlightens the mechanisms that underlie knee injuries, there are some issues that require author’s respond, particularly with regard to the method, the statistical analysis of the research data as well as the presentation and discussion of the results, before the study is considered for publication.

General comments

Method

• The authors should justify the choice of loads that they implemented for static loading of the knee joint as these could be considered light to mild in comparison to the loads that are developed in sporting activities such as during landing. This issue should be discussed more thoroughly and possibly introduced as one of the limitations of the present study.

R: This is a very good issue presented by the reviewer. The load type was performed to replicate the study performed by Chu et al. (2003). However, as the reviewer suggests, these loads may not have been sufficient for some participants. This has been added as a limitation of the study (p. 22 lines 452-456).

• The authors should also justify their choice to use the dominant leg (the one the participants use for kicking a ball) in their study as previous studies have demonstrated significant differences between the dominant and non-dominant leg regarding knee joint stability during unilateral landing (Ludwig et al 2017; Herrington 2011).

R: The use of the dominant leg has been used throughout the literature (Heebner et al., 2017; Jenkins et al., 2017; Kernozek et al., 2008). However, this does not discount the reviewer’s suggestion. This has been added as a limitation and framed as a topic for future work (p.22, lines 456-459).

• Given the complexity of the experiment, it is suggested that the researchers provide photographic material for the experimental set up and procedure.

R: Additional figure have been added to illustrate the experimental loading and landing protocols (p. 8 and 9).

Statistical analysis

The general sense that someone gets from reading this manuscript is data over-analysis. The authors performed several statistical tests in order to identify potential effect of static loading of the knee’s passive structures during landing. However, there is a mismatch (in my opinion) between the statistical tests that were performed, the presentation of the Results and the data listed or depicted in Tables and Graphs, respectively. More specifically:

• The authors performed a Two-Way (condition x muscle) ANOVA to compare average and maximum normalized EMG of each muscle involved during landing between pre and post loading conditions (the results are listed in Table 1); they also performed a 5 (muscles) x 2 (conditions) x 4 (time intervals) Two-Way ANOVA for average normalized EMG (the results are graphically presented in Figure 1). It is clear that in the latter case the authors attempted to investigate how muscles’ EMG behave over time during landing; however in Figure 1 the normalized EMG that was recorded before landing (200-0 and 100-0 ms) is also depicted. Why the authors did not include the pre-landing EMG’s in the statistical analysis? Their decision should be is justified and, in any case, they should consider modifying Figure 1, as in its current state it seems that both pre-landing and during landing EMG responses were also compared.

R: Thank you for finding these issues in the statistical analysis section. For clarity, the explanation of the data analysis for each dependent variable has been revised. When EMG analysis is conducted, the 200 ms before and 200 ms after landing are explicitly stated.

• Similarly, EMG muscles ratios have been compare with two different statistical tests. A muscle ratio x condition Two-Way ANOVA was used to compare average and maximum EMG ratios between the pre- and post-loading condition (the results are listed in Table 3 and 4) and again a 6 (muscle ratios) x 2 (conditions) x 5 (time intervals) Two-Way ANOVA for muscles EMG ratios (results are graphically presented in Figure 2). In this case (i) the authors did not report whether muscles EMG ratios were based on the average or the maximum EMG activity, (ii) they included 5 time intervals whilst the time intervals during landing were 4; which was the 5th time interval? Did the authors include in their analysis a pre-landing time interval (200-0 or 100-0 ms)? And if they do so, why they did not the same for average and maximum EMG activity of the muscles under investigation? (iii) an additional column was added in Figure 2 (the one that represents total EMG ratio for each time interval) but it should be clarified whether or not it was included in the statistical analysis. The authors should consider omitting the “total EMG ratio column” particularly if its scientific value is meaningless as it confuses the reader (see next comment).

R: To clarify the data and simplify the Results section, the ratio data have now been excluded from the data analysis.

• In both aforementioned cases the “overall” average and maximum EMG as well as the “overall” muscle EMG ratios is presented in Tables 2-4. In Figure 2 and Figure 3 a “total” EMG ratio column and a “combined” vertical ground reaction forces column (VGRF), respectively is also presented. The authors should justify whether (i) the “overall”, “total” or “combined” are terms that were used interchangeably, (ii) these terms represent data obtained at the pre-landing and during landing condition (which in my opinion make sense) or data obtained before loading and after loading of the passive knee structures. The legends in Tables 3 and 4 add more confusion to the reader as it is not clear whether the data listed in Tables represent “… average and maximal muscle ratios during landing phase between pre and post landing conditions” or “… loading conditions”. In any case the authors should consider omitting the “overall”, “total” or “combined” data, as in my opinion this information neither adds more scientific value to this research nor it is thoroughly discussed.

R: There is agreement with the reviewer that the “overall” and “total” data do not provide meaningful information. These have been removed from the figures and tables. The Figure legends and table legends have been updated to reflect a clearer reflection of the presented data.

• For VGRF the authors (i) reported the test that they used in statistical analysis, (ii) in the Results section they reported no statistical differences between conditions with regard to absolute peak and normalized peak forces but (iii) in the Discussion section they stated that “significant reductions in the average VGRF after static loading of the knee was present” (Page 20, Lines 429-430). This information was probably extracted from the graph in Figure 3 (?), but the authors neglected to report the level of statistical significances either in the text or in the graph.

R: These VGRF results have been included in the Results section of the revised manuscript (p. 17, lines 308-310). Additionally, the normalized forces have been omitted from the revised manuscript.

• The statistical tests used for maximum and minimum angular displacement, angular velocities and ROM comparisons between conditions were not reported.

R: These statistical tests are now included in the revised manuscript (p. 11, lines 225-227).

Discussion

The authors should discuss the limitations of the study and propose future research based on the findings and / or the methodological Inadequacies of the present study.

R: A section has been included to highlight the limitations and potential future studies related to these limitations (p. 22, lines 452-461).

Minor comments

Page 3, Line 47: The authors should consider replacing “… stress and strains which …” with “… stress and strains, which…”

R: This has been changed, as suggested by the reviewer.

Page 3, Line 61: The authors should clarify whether they are referring to the anatomical (malalignment of mechanical axes) or dynamic “… knee valgus …”

R: This point has been clarified to the dynamics of the movement.

Page 4, Line 68: The authors should consider replacing “…musculotendonous…” with “…musculotendinous…”

R: This spelling has been corrected throughout the document.

Page 6, Line 126: The authors should describe the technique used, if any, for identification of motor points and/or cite related references.

R: References for electrode placement have been provided in the methods (p. 6-7, lines 127-131).

Page 7, Lines 148-152: The authors should clarify the number of test trials that were actually included in data analysis as according to their protocol 10 test trials were required but eventually “… 5 sufficient trials…” were enough. Does this mean that the average of 5 trials out of the 10 was included in the data analysis and, if yes, which trials were discarded if more than 5 trials were recorded?

R: This has been clarified in the revised document.

Page 13, Line 276: The authors should replace (or not) the “… pre and post landing conditions” with “… pre and post loading conditions” in the legend of Table 3. The same applies for the legend of Table 4 (Page 14).

R: These changes have been made in the revised manuscript.

Page 13, Table 3: The authors should consider omitting the “overall” data (unless they explain and justify its use) since, in my opinion, it is confusing, not related with the initial hypothesis and eventually not discussed, although it appears that is the only data sets that present statistical significances. The same applies for the legend of Table 4 (Page 14).

R: As stated previously, this has been deleted from the figures, tables, and text of the manuscript.

Page 15, Line 307: The authors should consider replacing “… (p>0.071)” with “… (p<0.071)”

R: This has been changed.

Page 19, Lines 400-401: The authors should consider replacing “… overall knee rotation at landing” with “… overall knee flexion at landing”. They should also consider modifying Table 5 as in this Table the minimum and maximum, but no “overall”, displacement is listed.

R: These changes have been made to the manuscript.

Page 19, Line 407: The authors should consider replacing “… (Table 4)” should with “… (Table 5)” as the latter one display data values of joints angular displacements while the former displays data values of EMG muscle ratios.

R: This has been corrected in the manuscript.

Page 19, Lines 410-411: The authors should specify the negative values in both the text and in Tables 5 and 6.

R: This has been updated in the revised manuscript.

Figure 1: The units for NEMG should be added in the vertical axis

R: There are no units of normalized EMG. Rather, the data are provided with reference to the maximal EMG value of the respective muscle.

Figure 1-3: The author should use the same title in the horizontal axes (Time or Time intervals) in all Figures. The levels of statistical significance may also be indicated in a revised version of the manuscript provided that the proposed amendments have been taken into account.

Figure 3: The legend should be modified according to the other graphs for consistency. For example: Pre-loading � instead of Pre – black

R: These changes have been made in the revised manuscript.

Submitted filename: Response to Reviwers.docx

Click here for additional data file.

18 Dec 2019

PONE-D-19-17819R1

Static loading of the knee joint results in modified single leg landing biomechanics

PLOS ONE

Dear Dr. Olson,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

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Dragan Mirkov, Ph.D.

Academic Editor

PLOS ONE

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: All comments have been addressed

Reviewer #2: All comments have been addressed

**********

2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: No

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: No

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

Reviewer #2: Yes

**********

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: I still think that such a quality and correct methodology should have more subjects in the statistical analysis.

Reviewer #2: Review of the manuscript titled “Static loading of the knee joint results in modified single leg landing biomechanics”. Manuscript Number: PONE-D-19-17819.R1

The investigator of this study responded to all the comments and queries that were highlighted after reviewing the first version of the manuscript. However, there are several points that should be clarified, particularly with regard to the statistical analysis of the research data as well as the presentation and discussion of the results, before the study is considered for publication.

Abstract

Page 2, Lines 38-39: The authors should clarify the abbreviations aVGRF and NAEMG as they do not appear in the text

Methods

Pages 10-11, Lines 215-219: The author should consider re-writing the sentence following the suggestions presented below.

Statistical analysis

Page 11, Lines 224-234: The author should (i) clarify why Pre- vs. Post- comparisons, for angular displacement and velocity variables as well as for average and maximal forces, RFD and VGRF, are performed with a One-Way Analysis Of Variance and not a paired-t test, (ii) report the statistical test that he used for the analysis of the Force Timing-related data, (iii) consider to refer to the statistical tests used for each variable in the same order that it is presented in the Results section and discussed in the Discussion section. Alternatively, the statistical tests could be presented in groups (e.g. first the statistical test used for paired comparisons and then the ANOVA tests).

Results

Pages 12-16: The author should specify what the “pre- and post- landing condition” and the “pre- and post-loading condition” are. It appears that (i) the terms “pre- and post- landing” are used when the comparisons are intended to assess the effect of the pre-landing-induced feed forward mechanism on certain variables (e.g. muscle activity) and (ii) the terms “pre- and post-loading” when the comparisons aimed to assess the effect of passive loading of the knee on a particular variable.

An example: Table 2 presents comparisons between pre- and post-landing conditions for “Overall Average and Maximum NEMG” data. This is okay since the author stated (Pages 10-11, Lines 215-219) that such comparisons will provide information on the effect of the feed forward mechanism. However, it is not clear whether this represents the data that recorded before (pre-) or after (post-) passive loading of the knee. The author should (i) consider presenting pre- and post-loading and pre- and post-landing data in Tables or Figures as he did for the average NEMG data (Figure 3) and (ii) perform Two-Way ANOVAs for the maximum NEMG, as well as for all the other variables, as he did for average NEMG and VGRF.

Such clarifications should be made in the following cases, but generally this should be done whenever is deemed necessary.

Page 12, Line 250: In the text

Page 13, Line 253 and 255: In the text and the legend of Table 2. It should be clarified whether the data presented refers to the pre-loading or the post-loading condition

Page 13, Line 257: Consider replacing “…during drop landing conditions.” with “…during drop landing.”

Page 13, Line 263-272: The author should (i) clarify in the text the "conditions" that he is referred to and (ii) report in the legend of Figure 3 that the data is referred to both pre- and post-loading conditions.

Page 14, Line 275 and 279: In the text and legend of Table 3 for angular displacement data

Page 14, Line 282-286 and Page 15, Line 294: In the text and legend of Table 4 for angular velocity data

Page 15, Line 296-297 and 303-304: In the text and legend of Table 5 for joint ROM data

Page 15-16, Lines 306-310: In the text and legend (Page 16, Line 312-314) of Figure 4 for VGRF (How is it possible to record VGRF before landing?).

Page 16, Line 316: The author should consider defining in the Method section the Force Timing variable and report the statistical test that it was used for its analysis before presenting the related Results, as this variable appears for the first time in this section. Furthermore, Figure 4 presents maximum VGRF and not Force Timing data the values of which remain unspecified.

Page 12, Line 252: The author should consider replacing “Maximal” with “Maximum” and “EMG” with “NEMG” wherever is deemed appropriate.

Table 2 and Figure 3: The presentation of the values in Table 2 and Figure 3 for Average NEMG is inconsistent. The values presented in Table 2 are percentages of MVIE whilst the values presented in Figure 3 vary between 0-1.5 but remain unspecified. The authors should consider revising either the data in the Table or the data in the Figure.

Tables 3-5: The author should clarify the movement of ankle - dorsiflexion or plantarflexion - that is presented in the Tables and to explain the negative numbers.

Figure 4: The author should consider replacing the labels “Pre-landing” and “Post-landing” with “Pre-loading” and “Post-loading” that is presented within Figure 4.

Discussion

The discussion is very well written but the author must explain which conditions were compared. This is particularly important for the understanding of the discussion related to the Kinematic and Kinetic data.

**********

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Reviewer #1: No

Reviewer #2: Yes: Dimitris Mandalidis, Assistant Professor, School of Physical Education and Sports Science, National and Kapodistrian University of Athens

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Submitted filename: Reviewers comments for PONE-D-19-17819.R1.docx

Click here for additional data file.

15 Jan 2020

Abstract

Page 2, Lines 38-39: The authors should clarify the abbreviations aVGRF and NAEMG as they do not appear in the text

R: The abbreviations have been modified to agree with the abbreviations in the text.

Methods

Pages 10-11, Lines 215-219: The author should consider re-writing the sentence following the suggestions presented below.

R: please see the response after the following reviewer suggestion

Statistical analysis

Page 11, Lines 224-234: The author should (i) clarify why Pre- vs. Post- comparisons, for angular displacement and velocity variables as well as for average and maximal forces, RFD and VGRF, are performed with a One-Way Analysis Of Variance and not a paired-t test, (ii) report the statistical test that he used for the analysis of the Force Timing-related data, (iii) consider to refer to the statistical tests used for each variable in the same order that it is presented in the Results section and discussed in the Discussion section. Alternatively, the statistical tests could be presented in groups (e.g. first the statistical test used for paired comparisons and then the ANOVA tests).

R: It was believed that the ANOVA test was a valid statistical assessment of the values provided between the two conditions. In viewing the one-way ANOVA and paired-t-test options, both would provide the same outcome, thus the one-way ANOVA has been reported.

Results

Pages 12-16: The author should specify what the “pre- and post- landing condition” and the “pre- and post-loading condition” are. It appears that (i) the terms “pre- and post- landing” are used when the comparisons are intended to assess the effect of the pre-landing-induced feed forward mechanism on certain variables (e.g. muscle activity) and (ii) the terms “pre- and post-loading” when the comparisons aimed to assess the effect of passive loading of the knee on a particular variable.

R: The text has been edited to clarify pre-loading landing and post-loading landing conditions – these are the only “conditions”, as defined in the Methods section

An example: Table 2 presents comparisons between pre- and post-landing conditions for “Overall Average and Maximum NEMG” data. This is okay since the author stated (Pages 10-11, Lines 215-219) that such comparisons will provide information on the effect of the feed forward mechanism. However, it is not clear whether this represents the data that recorded before (pre-) or after (post-) passive loading of the knee. The author should (i) consider presenting pre- and post-loading and pre- and post-landing data in Tables or Figures as he did for the average NEMG data (Figure 3) and (ii) perform Two-Way ANOVAs for the maximum NEMG, as well as for all the other variables, as he did for average NEMG and VGRF.

Such clarifications should be made in the following cases, but generally this should be done whenever is deemed necessary.

R: All comparisons are between pre-loading landing and post-loading landing conditions. This nomenclature has been modified to reflect the clarity of the conditions in the text. The NEMG was evaluated 200 ms pre-landing and 200 ms post-landing, while the VGRF data were evaluated 200 ms post-landing in each condition. The data are presented in either figures or tables in the manuscript. The “overall average and maximal” values were removed for the first revision, so it is unclear what the reviewer is viewing.

Page 12, Line 250: In the text

R: the conditions have been defined in the Methods section for clarity

Page 13, Line 253 and 255: In the text and the legend of Table 2. It should be clarified whether the data presented refers to the pre-loading or the post-loading condition

R: this has been clarified

Page 13, Line 257: Consider replacing “…during drop landing conditions.” with “…during drop landing.”

R: this has been clarified

Page 13, Line 263-272: The author should (i) clarify in the text the "conditions" that he is referred to and (ii) report in the legend of Figure 3 that the data is referred to both pre- and post-loading conditions.

R: these have been clarified

Page 14, Line 275 and 279: In the text and legend of Table 3 for angular displacement data

R: this has been clarified

Page 14, Line 282-286 and Page 15, Line 294: In the text and legend of Table 4 for angular velocity data

R: this has been clarified

Page 15, Line 296-297 and 303-304: In the text and legend of Table 5 for joint ROM data

R: this has been clarified

Page 15-16, Lines 306-310: In the text and legend (Page 16, Line 312-314) of Figure 4 for VGRF (How is it possible to record VGRF before landing?).

R: it is not. It is not clear what the reviewer is reading, but there has been no indication in this manuscript that forces were recorded before landing.

Page 16, Line 316: The author should consider defining in the Method section the Force Timing variable and report the statistical test that it was used for its analysis before presenting the related Results, as this variable appears for the first time in this section. Furthermore, Figure 4 presents maximum VGRF and not Force Timing data the values of which remain unspecified.

R: this has been defined in the Methods section: “A 2-way ANOVA (condition x time interval) was used to assess average VGRF data during the landing phase, while one-way ANOVAs were used to compare maximal VGRF values between conditions”, p.11, lines 231-234. The caption of the section has been changed to clarify this variable assessment.

Page 12, Line 252: The author should consider replacing “Maximal” with “Maximum” and “EMG” with “NEMG” wherever is deemed appropriate.

R: this has been clarified

Table 2 and Figure 3: The presentation of the values in Table 2 and Figure 3 for Average NEMG is inconsistent. The values presented in Table 2 are percentages of MVIE whilst the values presented in Figure 3 vary between 0-1.5 but remain unspecified. The authors should consider revising either the data in the Table or the data in the Figure.

R: this has been clarified in Table 2

Tables 3-5: The author should clarify the movement of ankle - dorsiflexion or plantarflexion - that is presented in the Tables and to explain the negative numbers.

R: this has been clarified in the footnote for tables 3. Table 4 provides angular velocity data and the signs are indicative of positive and negative rates of movement, respectively. Table 5 represents ROM and does not require and explanation for the ankle

Figure 4: The author should consider replacing the labels “Pre-landing” and “Post-landing” with “Pre-loading” and “Post-loading” that is presented within Figure 4.

R: this has been modified.

Discussion

The discussion is very well written but the author must explain which conditions were compared. This is particularly important for the understanding of the discussion related to the Kinematic and Kinetic data.

R: the conditions have been clarified in the Discussion section

Submitted filename: Response to Reviewers 2 revision.docx

Click here for additional data file.

5 Feb 2020

Static loading of the knee joint results in modified single leg landing biomechanics

PONE-D-19-17819R2

Dear Dr. Olson,

We are pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it complies with all outstanding technical requirements.

Within one week, you will receive an e-mail containing information on the amendments required prior to publication. When all required modifications have been addressed, you will receive a formal acceptance letter and your manuscript will proceed to our production department and be scheduled for publication.

Shortly after the formal acceptance letter is sent, an invoice for payment will follow. To ensure an efficient production and billing process, please log into Editorial Manager at https://www.editorialmanager.com/pone/, click the "Update My Information" link at the top of the page, and update your user information. If you have any billing related questions, please contact our Author Billing department directly at authorbilling@plos.org.

If your institution or institutions have a press office, please notify them about your upcoming paper to enable them to help maximize its impact. If they will be preparing press materials for this manuscript, you must inform our press team as soon as possible and no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org.

With kind regards,

Dragan Mirkov, Ph.D.

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: (No Response)

Reviewer #2: (No Response)

**********

2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: (No Response)

Reviewer #2: No

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

Reviewer #2: Yes

**********

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: (No Response)

Reviewer #2: Review of the manuscript titled “Static loading of the knee joint results in modified single leg landing biomechanics”. (PONE-D-19-17819)

Dear Dr. Mirkov

I would like to thank you, once again, for giving me the opportunity to review the manuscript titled “Static loading of the knee joint results in modified single leg landing biomechanics”. This is undoubtedly a very important work the results of which will help both researchers and clinical therapists to better understand the mechanism of knee injuries. Unfortunately, many times the reviewer is obliged to identify ambiguities that are clear in the researcher's mind, only to improve the image of the manuscript to third parties and not to question the value of the work. This is often a demanding task and often frustrates the researcher as the project's publication is delayed.

The main concern of the last revision was the clarification of the terms "pre-landing" and "post-landing" conditions as well as the terms "pre-loading" and "post-loading" conditions which were used alternatively throughout the manuscript. Although it was clear to me that comparisons were made before and after passive knee loading, that is “pre- and post-loading” of the knee, this was not consistently reported (as in the case of ground reaction forces-GRF in both text and image), confusing the reader. However, the explanatory terms given by the author are sufficient and understandable. In the spirit of the above comments, the author should consider modifying the point that was addressed in the previous revision and although it was claimed that was modified it is not clear to me if this was actually happened. The point I am referring to is in the Abstract Page 2, Line 38: What "a" stands for in the abbreviation "aVGRF"? and in Line 39: What NAEMG stand for? Is it different from the NEMG that has been modified and used throughout the manuscript?

Furthermore the “Overall Average and Maximal NEMG” data that I was referring to, although it was removed from the Table in a previous modification of the manuscript, it remains as a subtitle of the entire paragraph that is dedicated to the results obtained from the analysis of EMG signals. Although its correctness/accuracy was not questioned the author should consider, in the final version of the manuscript, to modify the subtitle since this paragraph contains multiple comparison of the EMG data and not just the “Overall”.

Finally, the negative signs, in Table 3 have been clarified only for “Ankle flexion”. The negative signs for “Knee abduction” and “Hip abduction” have not yet been clarified (as proposed in the previous revision). Since the author clarify the negative signs in Table 4 for angular velocity (“…are indicative of positive and negative rates of movement, respectively”) he may consider adding the information as a footnote in the Table.

Sincerely

**********

7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No

Reviewer #2: Yes: Dimitris Mandalidis, Assistant Professor, Department of Physical Education and Sports Science, National and Kapodistrian University of Athens

7 Feb 2020

PONE-D-19-17819R2

Static loading of the knee joint results in modified single leg landing biomechanics

Dear Dr. Olson:

I am pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department.

If your institution or institutions have a press office, please notify them about your upcoming paper at this point, to enable them to help maximize its impact. If they will be preparing press materials for this manuscript, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information please contact onepress@plos.org.

For any other questions or concerns, please email plosone@plos.org.

Thank you for submitting your work to PLOS ONE.

With kind regards,

PLOS ONE Editorial Office Staff

on behalf of

Dr. Dragan Mirkov

Academic Editor

PLOS ONE