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Core E - Human Translational Studies

Core Director: Susan Harkema, Ph.D.




Personnel: Christie Ferreira, M. S.

 

The Human Locomotion Research Laboratory has been operational for 12 years, most recently at the University of Louisville and Frazier Rehab Institute, for 4 years.  Currently, the Human Locomotion Research Team routinely performs the following training paradigms, measurements, and corresponding data analyses for individuals after spinal cord injury:

    Training Paradigms
  1. Activity-based retraining of standing and stepping using the Therastride body weight supported treadmill system and manual assistance.
  2. Functional electrical stimulation supine and in combination with stand retraining.
    Experimental Data Acquisition
  1. Surface and fine wire electromyography (EMG) of trunk, arms and legs, kinetic and kinematic data during rest, standing and stepping.  
  2. Reflex modulation of leg muscles during rest, standing and stepping.
  3. Brain Motor Control Assessment and Transmagnetic motor evoked potentials during standardized manuevers that provide neurophysiological data that quantifies neural status.
  4. Pulmonary function parameters at rest.
  5. Cardiovascular parameters during rest, orthostatic stress, standing and walking.
  6. Standard behavioral and clinical measures of function including the 6 min walk, American Spinal Injury Association Injury assessment, among others.
  7. Magnetic Resonance Imaging of muscle for the assessment of muscle mass and fat content in lower extremities.
  8. Dual-energy X-ray absorptiometry (DEXA) scanning for the assessment of bone minenal density.

 

Core personnel.  

  • Dr. Harkema is the Core Director and Director of the NeuroRecovery Network and oversees 7 specialized centers that provide standardized activity-based rehabilitation care based on current scientific and clinical evidence for people with spinal cord injury and other selected neurological disorders.  
  • Ms. Christie Ferriera, Research Associate, has over 7 years of experience in all aspects of data acquisition and analyses, research participant recruitment and screening, and human use procedures.  Other members of the Human Locomotion Research team will be available and can participate as needed, and deemed necessary.  Personnel experienced in the research protocols and training paradigms will be available and allocated depending on the needs of the project from the disciplines of physiatry, physical therapy, engineering, and neurophysiology.   Highly skilled training teams in the activity-dependent rehabilitation paradigms are also available for the users.

 

Operating procedures.  

  • Access:  Before Users are given access to the facility they will provide a copy of HIPPA and CITI certification.  A written proposal of the research aims and design will be submitted to the Core Director and implemented in the order or request.  For peer-reviewed funded studies, Dr. Harkema and Ms. Ferreira will identify the resources and personnel required and develop an experimental and training schedule.  Core research team members will be selected to support the studies as appropriate with the intent to train the investigators to implement their protocols independently.  In case experiments are not funded by peer-reviewed grants, the scientific value of the experiments will be evaluated for approval by the Core director in consultation with the Steering Committee.  Upon approval, the investigator will follow the standard outlined for funded investigations.  If projects exceed capacity, priority will be given to funded studies and then based on rating of scientific value as determined by the Steering Committee.  Those supported by the pilot studies will be given funded priority.
  • Quality control:   An experienced Core F research team member will be assigned as an advocate for each research participant with a neurological injury involved in the study.  However, Core F personnel will continuously monitor the progress of the studies to ensure compliance with standard operating procedures and Institutional Review Board requirements.  Investigators will be required to attend monthly team meetings that focus on the research participants from all studies issues and progress.  Users will be appropriately trained, when necessary.     
  • Assistance:  Dr. Harkema and Ms. Ferreira will evaluate the needs of each investigator and provide the personnel required to achieve the research aim.  These requirements will vary depending on the level of experience of the investigator, the research aim and whether assessments alone or assessments and training interventions are required.  
  • Scheduling and Investigator support:  Users will schedule training sessions by contacting Ms. Ferreira after their study has been approved.  Scheduling for the use of the Core and for training is done during a consultation with Ms. Ferreira.  She has the experience to know the resources and recruitment procedures required for specific protocols and assessments.  The Human Locomotion Research Center maintains a calendar that is available to all users that includes type of intervention and assessment, location, time and duration of research event, lead investigator, research participant identification number, and personnel required.  Typically, users will observe and participate when appropriate in required assessments and/or training interventions.   Core personnel and designated HLRC research staff are constantly available for consultation. 

 

Core_F_Fig_1a.jpg

Fig. 1a. Acquisition of EMG, Kinetic and Kinematic Data. Top panel: general acquisition set-up. Bottom left panel: acquisition of kinematics and EMG during stepping. Bottom right: electrode placement for EMG acquisition, and motion analysis marker set-up.

 

Core_F_Fig_1b.jpg 

Fig. 1b. Incomplete SCI subject developed EMG activity during stepping in both ankle and knee muscles, which was absent prior to locomotor training.  EMG and joint angles are plotted over two steps during stepping using BWST at 27% BWL and 1.5 m/s before and after locomotor training. 

 

1. Training Paradigms

a. Activity-based retraining of standing and stepping using BWST. During the session, subjects are placed on the treadmill in an upright position and suspended in a harness by an overhead pulley at the maximum load at which knee buckling and trunk collapse can be avoided.  A trainer is positioned behind the subject to aid in pelvis and trunk stabilization, as well as appropriate weight shifting and hip rotation during the step cycle.  The trainer ensures that the trunk and pelvis are not flexed or hyper-extended during stepping.  Trainers positioned at each limb provide manual assistance using a customized technique developed by this research team that facilitates knee extension during stance and knee flexion and toe clearance during swing.  Trainers promote knee extension by applying gentle pressure at the tibial tuberosity and stimulation of the patellar tendon.  They promote knee flexion and toe clearance by applying a gentle force at the medial hamstrings tendon (Fig. 30).  Manual assistance at the trunk-pelvis and at the legs is used only when needed.  During the session, the treadmill speed is adjusted to promote the best stepping pattern at the given body weight load (BWL).  Speeds are maintained within a normal walking speed range (0.89-1.34 m/s).  Body Weight Support (BWS) is continuously reduced over the course of the training sessions as the subjects increase their ability to bear weight on the lower limbs.  Manual assistance is reduced with independence of stepping.

For stand training, a trainer is positioned behind the subject to aid in pelvis and trunk stabilization. Two leg trainers aid in knee extension by pushing on the patellar tendon repeatedly to facilitate knee extension and push on the medial hamstring (behind the knee) to prevent hyperextension.  The overall goal is to obtain a stable control during stance (and limb control), which resembles normal standing and to practice the movement repetitively. This is determined by adjusting and ultimately minimizing BWS and minimizing manual assistance.

b. Functional electrical stimulation supine and in combination with stand retraining. Electrical stimulation is applied via bifurcated leads and self-adhesive reusable surface electrodes.  The electrodes are applied over the motor points (both legs) on the following muscles: quadriceps (QUAD), biceps femoris (BF), medial gastrocnemius (MG), and anterior tibialis (TA).  Two electrodes are used for each muscle.  Two EMPI “Respond Select Neuromuscular Stimulators” (Empi Inc., St. Paul, MN) are used to induce the electrical stimulation.  Electrical stimulation is applied with symmetrical biphasic pulses of 300 µs at 35 Hz delivered across a 1000 ohm load, over a duty cycle of 11 seconds on, 60 seconds off, with overlap during each contraction between the upper and lower leg.  Therefore, MG and TA, muscles will be contracted first for 4 seconds. The BF and QUAD will be contracted next for 7 seconds, while the other musculature is still being stimulated (11 second total). This is followed by 60 seconds of rest (no stimulation).  This timing and phasing of contractions was selected to promote the muscle groups to contract and relax alternately in an overlapping fashion. Since maximum FES-inducible contraction is required for the venous pump to be effective, subjects are acclimated to the FES prior to training to find the maximum tolerable level of stimulation for each and apply that during the training.  Stimulation starts at the threshold level for minimal contraction and will advance to the maximum contraction over 2 seconds. We have used Functional electrical stimulation as a training paradigm in combination with stand retraining mentioned above (Fig. 30).

 

Core_F_Fig_2.jpg 

Figure 2:  Step Training with BWST (Left panel). Trainers are position behind the subject and at the legs to provide manual assistance during the session. Functional electrical stimulation with Stand retraining (Right panel): Stimulating electrodes asre placed on flexors and extensors to promote co-activation of knee and ankle muscles, while trainers provide manual assistance to promote knee extension.

 

2. Experimental Data Acquisition.

a. EMG, Kinetic and Kinematic Data Acquisition. A 24-channel hardwired system (Konigsberg Instruments, Pasadena, CA) is used to acquire footswitch and EMG analog signals.  An EMG amplifier (bandpass filter 0.1 kiloHertz (kHz) to 1 kHz) coupled to a pulse interval modulator relays data to decoding electronics.  Data is digitally sampled by an on-line analog to digital (A-D) conversion system (Labview, National Instruments, Austin, TX) at 1000 Hertz (Hz) and processed by customized Labview acquisition software.  BWS is measured using a force transducer attached in series with the suspension apparatus.  BWS is acquired by Labview A-D and processed by customized software (Fig. 31,32).

Limb kinematics including hip, knee and ankle angles is acquired using high speed passive marker motion capture (Motion Analysis, Santa Rosa, CA), a real-time capture and display system.  It is a six-degree-of-freedom true 3D motion capture and analysis system that tracks x, y, and z coordinates and orientation angles concurrently.  Our lab has designed a marker system to prevent interference of the trainers or harness with the view of the markers by the cameras.

We measure individual ground reaction forces (GRF) using Fscan (TEKSCAN, Boston, MA) shoe-insole pressure sensors, and also capture the GRF during multiple footstrikes, dynamic weight transfer and local pressure concentrations during standing using HR Mat equipment and software (TEKSCAN, Boston, MA).  Calibration is verified by comparing signals to those obtained from a force platform (Kistler, Amherst, NY) with non-disabled individuals matched by body weight and shoe size to SCI subjects since the SCI subjects will be incapable of walking overground.  

b. Reflex modulation Acquisition. Changes in spinal reflexes are generally used as a measure of motorneuron excitability and to probe spinal excitability and pathology.  The H reflex is generally accepted as an indicator of the number of motor neurons recruited during the stimulation.  After spinal cord injury, the amplitude of spinal reflexes increases, and continues to do so over time.  We can record the soleus H-reflex while the individual with SCI is resting in a supine or sitted position, standing and/or stepping. The H-reflex is evoked by monopolar electrical stimulation of the posterior tibial nerve at the popliteal fossa. In total approximately 20 reflexes are recorded in every trial to generate a recruitment curve. Recordings are taken with stimuli of 80%, 90%, and 100% of maximum stimulus from the soleus muscle.  Each stimulation is a 1-ms monopolar electrical stimulation pulse, generated by a constant current stimulator (DS7A, Digitimer, UK) sent to an on-line analog to digital (A-D) conversion system (Labview, National Instruments, Austin, TX) at 1000 Hertz (Hz) and processed by customized Labview acquisition software. A monopolar electrode placed over the posterior tibial nerve at the popliteal fossa is used to evoke the H-reflex, while a bipolar electrode over the soleus muscle is used to record the reflex. The indifferent electrode is placed above the patella. The stimulating site is chosen as the one that the H-reflex is observed first on the oscilloscope without an M-wave being present; this is based on recruitment and conduction velocities of group Ιa muscle afferents and of motor axons. Similarly we have the ability to record other reflexes at rest or during standing and stepping with BWST.

We have published data on the modulation of H-reflex during stepping in spinal cord injured and non-disabled individuals. This work was done with a collaborating investigator at CUNY in NY and demonstrates the functionality of the core. We collected data on ten non-disabled individuals and nine individuals with spinal cord injury. We found that in non-disabled individuals the H-reflex modulation remains unchanged at different levels of BWS. A similar H-reflex modulation pattern was observed in three out of nine SCI subjects, while in the remaining subjects the soleus H-reflex was not progressively enhanced from early to late stance and was not depressed during the swing phase {knikou2008} (Fig. 3).

 

Core_F_Fig_3.jpg

Fig. 3. Soleus H-reflex modulation during stepping with BWST (A-B). Overall amplitude of the soleus H-reflex and M-wave normalized to the maximal M-wave during stepping at 0%, 25% and 50% BWS in non-disabled subjects. C. Soleus H-reflex, M-wave, and soleus background activity (% of Mmax) are indicated for AIS-D during 40% BWS at 0.88 m/s. (D) . Overall amplitude of the soleus H-reflex and M-wave during treadmill walking in SCI subjects at BWS levels that varied from 20 to 60%. Knikou2008

c.  Brain Motor Control Assessment and Transmagnetic Motor Evoked Potentials. The Brain Motor Control Assessment (BMCA) is a protocol of volitional and reflex motor tasks rigorously carried out with the subject in the supine position using published standards for administration.  Following standard skin preparation, pairs of surface EMG (sEMG) electrodes spaced 2 cm apart are placed over the muscle bellies of the right and left upper trapezius (UT), biceps brachi (BB), triceps brachi (TB), wrist extensors (WE), wrist flexors (WF), adductor policis brevis (APB), abductor digiti quinti (ADQ), rectus abdominus (RA) (para-umbilical), quadriceps (QD), adductor femoris (Add), hamstrings (H), tibialis anterior (TA), soleus (Sol), extensor digitorum brevis (EDB), and abductor hallicus (AH).  The protocol begins with 5 minutes of relaxation followed by three repetitions each of: reinforcement maneuvers (deep breath, neck flexion, shoulder shrug); voluntary movements of arms and legs; passive movements of arms and legs; manual ankle clonus elicitation; suppression of plantar stimulation cutaneomuscular reflex response.  Surface EMG (sEMG) signals are recorded on a 32-channel Eclipse Neurological Workstation (AXON Systems, Inc.) with a sampling rate of 2 KHz per channel and a bandpass of 30 Hz to 2 KHz.  

Transmagnetic Motor Evoked Potentials are elicited through transcranial magnetic stimulation of the motor cortex as single or paired-pulses (15 ms inter-pulse interval) delivered through a flat coil for upper-limb responses and a dual-cone coil for lower-limb responses.  The placement for the coils is centered over the scalp vertex as determined by the 10-20 Internation EEG placement scheme. The responses, called motor evoked potentials (MEPs), are elicited by progressively increasing stimuli delivered with at least 5 seconds.  transcranial magnetic stimulation is provided by two Magstim200 devices joined through a Bistem unit, each producing a 100 μsec pulse of up to 2 Tesla intensity (Fig. 34).  Beginning with single pulses, intensities are increased from 30% of maximum in 10% output increments until all target (upper or lower-limb depending on the coil) muscles show responses or maximum stimulus intensity is reached.  

d.  Pulmonary Function.  Spirometric parameters of forced vital capacity (FVC) and forced expiratory volume in one second (FEV1), maximum inspiratory pressure (MIP) and maximum expiratory pressure (MEP) are acquired while the participant is in a sitted position. Testing is performed using a preVentTM pneumotach with BreezeSuite System under force after a maximal inhalation. To measure FVC, the participant inhales maximally and then exhales as rapidly and as completely as possible. FEV1 is the volume of air exhaled in the first second repeated three times.  At least one minute of rest is allowed between each effort, and at least five minutes between each group of three efforts. Maximal Inspiratory Pressure (MIP) is the highest atmospheric pressure developed during inspiration against an occluded airway and Maximal Expiratory Pressure (MEP) is the highest pressure developed during expiration against an occluded airway. MIP and MEP values provide standard non-invasive indices of inspiratory and expiratory muscle function.  

e.  Cardiovascular Function. Continuous arterial blood pressure is acquired from a finger cuff placed around the left middle/index/thumb (Portapres Model-2; Finapres Medical System). The left hand is placed in the arm sling and kept at the level of the heart throughout the study. Manual arterial blood pressure measurements are taken at the beginning of the supine control period and at the end of the recovery period with a digital blood pressure measurement device. A three lead ECG (lead II; ML132, ADInstruments) is used for ECG monitoring. The rib cage and abdomen respiratory rates are acquired using an Inductotrace inductive plethysmograph (Inductotrace, Ambulatory Monitoring). Tilt angle can be acquired from an accelerometer (Applied Geomechanics) mounted on the top of the bed.

 

Core_F_Bone_Scan.jpg

f. Standard behavioral and clinical measures. Standard behavioral and clinical measurements will include assessment of muscle tone (Modified Ashworth), clonus, tendon reflexes, grip strength, American Spinal Cord Injury Association - Impairment Scale (AIS), independence, balance and functional mobility.  The Spinal Cord Independence Measure (SCIM) will assess abilities such as self-care, respiration and sphincter management and mobility. We will utilize the Berg, Tinetti, and Functional Reach Tests to assess the subject’s balance.  The Berg utilizes a 5 point ordinal scale to quantify a patient’s level of balance in 14 activities.  The Modified Functional Reach Test is based on the Functional Reach Test (FRT).  The FRT is a well established test of dynamic balance in the forward direction, that is sensitive to change and has demonstrated good reliability and validity for primarily the elderly individual.  The Tinetti is used to predict a patient’s risk of falls by objectifying gait and balance while the subject performs specific tasks based on a three-point scale. Functional mobility will be assessed using the WISCI scale during a 10-m walk, GAITRite®, and the 6-min walk test with the SCI-FAI.  The Walking Index for Spinal Cord Injury (WISCI) scale is a valid and reliable scale used to assess walking ability and improvements occurring as the result of treatment interventions.  The GAITRite® System will analyze footfall pressure distribution and the objective parameters of gait. The 6 min walk with Spinal Cord Injury – Functional Ambulation Inventory (SCI-FAI) will measure walking endurance and walking function.  

g.  Magnetic Resonance Imaging.  Through our partnership and collaborations with Jewish Hospital and St. Mary’s Healthcare we have open access to a 1.5 and 3.0 Tesla GE Signa MRI Systems. We acquire cross sectional areas of each lower limb (the origin of the iliospsoas to the insertion of achilles tendon) before and after the training interventions. Participants lie prone with their knees braced at full extension and their ankles in neutral position to limit muscle spasms. Saline bags are placed adjacent to the legs to increase the signal to noise ratio. We collect coronal-plane images to identify landmarks and axial-plane images at 10 mm intervals with no interslice gap to analyze muscle volumes. T1 weighted images with the following parameters are used to maximize soft tissue resolution: TR: 550 ms; TE: 20 ms; number of excitations: 2; and matrix: 256x256.

h.  Dual-energy X-ray absorptiometry. Dexa scans are obtained through the same partnership with Jewish Hospital and St. Mary’s Healthcare  as described above. The Lunar scanner (Lunar Inc., Madison, WI) uses a pencil beam X-ray to measure total body BMD and regional body components including the femoral neck, lumbar, proximal tibia and distal femur, and total body.  Participants are scanned in a supine position on a padded densiometer tabletop using two different levels of energy (40 and 70 Ke V) passing the patient below while the differential absorption is measured above. The ratio of absorption between the two radiographs of different energies is linearly related to the fat tissue in the soft tissue compartment. DEXA provides a three-compartment partition of the body: Bone mineral density, fat mass and fat free mass.

We have an on going collaboration with the Kessler Research Foundation in which we are investigating the basic musculoskeletal gains associated with stand training with electrical stimulation for individuals after an SCI compared to electrical stimualtion alone or stand training alone. Also, the project examines how these musculoskeletal gains affect neural and muscle activation during standing and stepping. This will identify a better approach for rehabilitation therapies for people with sub acute to chronic SCI. Preliminary results show that after 60 sessions of training, the stand training alone group loss bone across all sites measured about the knee, the electrical stimulation group gained bone, but the greatest increase in bone occurred for the combination group, stand training with electrical stimulation (Fig. 4). Similar results were established with the muscle volume (Fig. 4) in the lower limbs, for the stand alone group there was a loss in muscle volume and a gain in adipose tissue, for the electrical stimulation group there was a gain in muscle volume and adipose tissue, but for the combination training there was a gain muscle volume and adipose tissue.  Our data suggests that combination of loading and electrical stimulation has the greater effect on the musculoskeletal tissue than just the electrical stimulation alone or the loading alone.

 

Core_F_Fig_4.jpg

Fig. 4:. In stand training alone group, the % BMD decreased for all sites. For electrical stimulation alone group there was a increase in %BMD, but a greater increase in %BMD was seen in the stand training with electrical stimulation group. Percent change data from post to pre training for Muscle, Intra Muscular Fat and Subcutaneous Fat for five individuals.  ES + Stand training increases muscle mass and reduces intra-muscular fat and subcutaneous fat.

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