Participants and 84 years (m=69,92; std=4,14), which is

Participants

To compare the muscle properties and fatigue resistance of the knee extensor muscles with the possible influences of physical activity and protein intake we include two separate groups of participants. The first group consists of 60 physical active elderlies aged between 65 and 84 years (m=69,92; std=4,14), which is part of a larger ongoing study ProWalk at Radboud Nijmegen Medical Centre. The second group consists of 20 sedentary elderlies who do not meet the norm of physical activity of the age 55+ years (Minimal a half hour physical activity at the intensity of 3 to 5 MET on at least 5 of the 7 days, or three times a week heavy physical activity for at least 20 min.) aged between 65 and 70 years old (m= ; std= )IK1 . Exclusion criteria are; health problems as heart and vascular diseases, neurological diseases, knee and hip problems. Participants signed informed consent. This study was approved in Radboud University Nijmegen Medical Centre by the medical ethical committee, and approved in Amsterdam by the ethics committee.

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General design

The muscle properties of the physically active elderly were measured in one occasion at Radboud University Nijmegen Medical Centre, and the muscle properties of the sedentary elderly in one occasion at Vrije University Amsterdam. The questionnaire about nutritional intake and physical activity level were taken at the same occasion. In advance and on the occasion participants were informed about the testing procedure. All tests were conducted under standard conditions. The room temperature was maintained around 21?C.

Muscle properties

Participants were not allowed to perform strenuous exercise 24-hour before the measurements. All measurements of the muscle properties were performed on a custom-build dynamometer with the knee flexed at 60° degrees (0° corresponds to full knee extension) and the hip at 70° flexion (0° corresponds to full hip extension). To avoid changes in hip and knee angle during the measurements the participants will be firmly strapped to the dynamometer with a belt around the hip and trunk. The participants’ ankle of their preferred leg will be tightly strapped to a force transducer with a padded cuff 2 cm above the ankle joint. All force signals were sampled at 1000 Hz and low pass filtered at 50 Hz. The distance between lateral epicondyle and fixed point on the force transducer will be used to calculate the moment arm.  All force signals were sampled at 1000 Hz and low pass filtered at 50 Hz.

Force measurements

Maximal isometric force recordings were made from voluntary contractions. Participants performed three maximal voluntary contractions (MVCs) of three to four seconds with a rest of two minutes in between. When the last MVC was 10% higher than the previous MVC, another MVC was performed, with a maximum of five MVCs. Participants will be strongly encouraged to exceed their previous maximal value. The values were displayed on a computer screen to confirm the subjects’ achievements throughout the test (Horstman et al. 2010).

Electrical stimulation

After determining the MVC, two self- adhesive surface electrodes (Schwa-medico) of 80 mm x 130 mm were positioned over the proximal and distal part of the quadriceps femoris muscle. Prior applying the surface electrodes the skin was shaved and cleaned with Orphilon Chlorhexidine 0,5%mV in alcohol 70% V/V. A computer controlled electrical stimulation current with a pulse of 200 ?s from a constant current stimulator (model?) starting at 40 mA and build up in maximal seven steps till a force was generated approximately 30-40% of the participant MVC.  Assuming 30-40% of the muscle mass was stimulated. All the stimulation intensities were conducted on 50 Hz for one second. The participants were instructed to relax the muscle as much as possible. The current intensity determined by 30-40% of the participants MVC was maintained throughout the rest of the tests determining the contractile properties (Mulder et al. 2006).

Voluntary activation capacityIK2 

 

Force-frequency relationship, force development and relaxation

First the isometric force-frequency relationship was obtained using 1-s contractions at different imposed stimulation frequencies (1, 10, 30, 50, and 100 Hz) separated by one-minute rest.  To obtain the maximal rate of isometric force development an 80-ms contraction at 300 Hz was imposed (Gerrits et al. 1999; Mulder et al. 2008). The half relaxation time (½RT) was assessed from the time needed to the decline from 50% to 25% of the peak force of the 100 Hz stimulation (De Ruiter, C.J., and A. De Haan (2000).

Fatigue resistance

The resistance of fatigue was assessed by activating the knee extensor muscles repetitively using 50 Hz bursts of one second duration with one second rest in between for a period of two minutes (60 contractions) (Voorn et al 2014).  

Assessment of protein intake and physical activity

At the same occasion participants were assessed on the nutritional intake with the Food Frequency Questionnaire (FFQ) (Faure et al. 2017), and the level of physical activity with the Short QUestionnaire to ASsess Health-enhancing physical activity (SQUASH) (de Hollander et al. 2012).

Food Frequency Questionnaire (FFQ)

The FFQ is a self-administered, paper-based, semi quantitative 110-item food-frequency questionnaire. For each FFQ item, protein intake was estimated based on the German Food code and Nutrient Database (BLS 3.02) by using the nutritional software PRODI (version?). IK3 For each food item participants could choose their usual rate of consumption, and the servings from common portions (Faure et al. 2017).

The Short QUestionnaire to ASsess health-enhancing physical activity (SQUASH)

The SQUASH has been designed to measure the habitual activity level in general. It contains questions about multiple activities referring to a normal week in the past 12 months divided by physical activity patterns in the house, travelling to work, and physical activity at work. The intensity of physical activity is divided by light, moderate, and vigorous (Wendel-Vos et al. 2003).

Data analysis

Off-line analysis of force signals was performed using custom-made (MATLAB) programs. Force was converted to torque with the moments arm calculated from the rotation point in the knee to the force transducer.

Voluntary activation capacityIK4 

 

Torque-frequency relationship

For each experimental session the peak torque of the 1, 10, 30, 50, and 100 Hz were expressed relative to the maximal torque reached during the 100 Hz tetanus (Mulder et al. 2008).

Torque development and relaxation

The torque development was quantified by assessing the time to peak tension from the start of the evoked contraction of the 300 Hz stimulation (TPT300). The start of the contraction was defined as the instant the first puls of the 80 ms train was delivered normalized to the maximum torque reached at the 100 Hz tetanus (Mulder et al. 2008).  For the maximal rate of relaxation, the late half-relaxion time is used (½RT). This is the time needed for force to fall from 50% to 25% of its maximal value reached at the 100 Hz stimulation (De Ruiter, C.J., and A. De Haan 2000). The Force Oscillation Amplitude (FOA) relative to the mean force was assessed during the 10 Hz stimulation using:

where dF is the average amplitude over three consecutive oscillations in the tetanus after the tension reached its peak ad Fmean is the mean force during that time (Gerrits et al. 1999).

Fatigue resistance

Fatigue resistance was expressed as the toque at the end of the fatigue protocol divided by that at the start of the protocol (Voorn et al 2014).

FFQ

For each FFQ item, protein intake was estimated based on the German Food code and Nutrient Database (BLS 3.02) by using the nutritional software PRODI (version?). Protein intake was estimate in units of grams per day, respectively (Faure et al. 2017).

SQUASH

Based on reported effort in the questionnaire, activities were given an intensity score ranking from 1 to 9. Household work and activities at work or school were restructured for intensity. Therefore, for these items we used a basic intensity score of 2 and 5 for light and intense activities, respectively. Total minutes of activity were calculated for each question by multiplying frequency (days/week) by duration (min/day). Activity scores for separate questions were calculated by multiplying total minutes of activity by the intensity score. The total activity score was calculated by taking the sum of the activity scores for separate questions.

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