This study was funded by the Coordination for the Improvement of Higher Education Personnel of Brazil (CAPES- #1800789).

The study was approved by the Research Ethics Committee of the University of Campinas (UNICAMP) under Protocol Number 2479761. Written informed consent was obtained in-person from all study participants. Sixteen participants of both sexes were recruited from the community of the metropolitan region of Campinas by searching the database of the Applied Kinesiology Laboratory (AKL). The inclusion criteria were 1) individuals aged 60 and older; 2) ability to answer the questionnaires used in this study; 3) ability to walk independently without difficulty; 4) availability to participate in the intervention during 3 months with at least 90% attendance. The exclusion criteria were 1) severe cardiovascular or pulmonary diseases; 2) orthopedic diseases that prevented performing daily activities; and 3) diseases that increased the risk of falls such as Parkinson&#39;s and Alzheimer&#39;s. All assessments were performed by researchers and professional physical trainers from the AKL.
1. Assessment
<ol>
	<li>Assess physical fitness using the following functional tests: Illinois agility, five times sit-to-stand, timed up-and-go, walking usual speed, and one-leg stand. Perform all tests before and after training. 
	NOTE: Here, the participants repeated each test 2x, and the best result was used.
	<ol>
		<li>Illinois agility test: Have the participants move through obstacles in multiple directions.
		<ol>
			<li>The length of the course is 10 m and the width is 5 m. Place a marking cone in each corner of the course: a start and finish cone at one end, 5 m apart; two at the turning points on the other end of the course, 5 m apart; and four cones in the center of the course spaced 3.3 m apart from each other.</li>
			<li>Have the participants jog in a straight line from the start line to the first turning point and then in the opposite direction towards the first center cone. They should weave back and forth through the four center cones from one end of the course to the other. Then, they should run diagonally from the first center cone to the second turning point located on the far right and then in the opposite direction towards the finish line14.</li>
		</ol>
		</li>
		<li>Five times sit-to-stand test: Have the participants rise from a chair 5x as quickly as possible with the arms crossed in front of the body, lifting the hips from the chair when the timer is started. The participants should stand up completely by extending the knees and sit down completely by maintaining the back against the chair15. Stop the timer after the participant sits down for the fifth time.</li>
		<li>Timed up-and-go (TUG) test: Have the participants get up from a chair without pushing up with their arms, walk as quickly as possible along a 3 m line demarcated on the floor upon hearing the command &#34;Go,&#34; return to the same position, and sit down on the chair again. Timing starts as the participants get up from their chair and stopped when the participant sits down16.</li>
		<li>Walking usual speed test: Have the participants walk 10 m at their usual gait. Start the timer when a foot reaches the 1 m line and stop when a foot reaches the 11 m line. Do not include the first and last 1 m stretches in the analysis because they are used to measure acceleration and deceleration, respectively17. Present the results as velocity in m/s (Figure 2).</li>
		<li>One-leg stand test: Have the participants stand on the foot from the dominant leg while flexing the contralateral knee at 90&#176;, folding the arms across the chest, and maintaining the head straight. Have the participants lift one foot from the floor when the timer is activated. Stop the timer when the foot touches the floor again. The maximum balance time is 30 seconds for each side. Test both legs18.</li>
	</ol>
	</li>
</ol>
2. Training program design
<ol>
	<li>Have the participants perform agility training for 12 min, including four 3 min sets with a 1 min rest between each set (i.e., the first set starts with sequence 1). Each set is composed of four 30 s subsets, a 15 s rest between subsets (Figure 3), and 1 min rest between sets.
	<ol>
		<li>In addition to the four exercise sequences, include a 10 min warm-up and 5 min cool-down, using whole-body low-intensity stretching. 
		NOTE: During this program, each participant trained 2x per week for 14 weeks.</li>
	</ol>
	</li>
	<li>Increase the level of difficulty by augmenting exercise complexity and volume, depending on individual adaptation to the program. The level of adaptation dictates the level of increase in the velocity of the drills.
	<ol>
		<li>Increase training complexity after 6 weeks of training with four sequences of coordination. After a period of adaptation, substitute sequences 2 and 3 for more challenging drills.</li>
		<li>Increase the training time after 10 weeks of training. Start with 12 min of exercise (four 3 min sets) and end with 15 min (five 3 min sets). 
		NOTE: At the end of each 30 s subset, the time taken to perform the activity is recorded. This procedure allows comparison of the distance travelled by each participant during each session of the training program.</li>
	</ol>
	</li>
</ol>
3. Agility ladder
<ol>
	<li>Perform the exercises using a ladder with a length of 4.8 m and 12 transverse rungs forming 12 squares (areas delimited by the rungs) measuring 40 cm 40 cm (Figure 1). Instruct the study subjects to step in the squares and not on the rungs.</li>
</ol>
4. Familiarization period
<ol>
	<li>Perform four sequences of the training program (1, 2A, 3A, and 4) during the 2 week familiarization period. Exercise sessions were spaced at least 48 h apart, and there was no time limit to complete the sequences in each session. Instruct the participants to practice these sequences to perform as well as possible during the training period.</li>
</ol>
5. Training program
NOTE: All drills start with both feet together on the start line.
<ol>
	<li>Sequence 1: Have participants place their left foot in the first square, the right foot in the second square, and so forth (Figure 4). Each sequence should be performed in both directions on the ladder for 30 s.</li>
	<li>Sequence 2A: Have participants position their right foot in the first square, then position the toes of the left foot in the first square, move the left foot to the second square, place the toes of the right foot on the second square, and so on (Figure 5).</li>
	<li>Sequence 2B: Increase the level of difficulty depending on individual adaptation to sequence 2A (Figure 6) by having participants move through the ladder in small jumps, with toe-tapping movements.
	<ol>
		<li>Have participants jump to the first square with both feet together such that the sole of the right foot touches the ground while the toes of the left foot touch the ground.</li>
		<li>Then, have them jump to the second square with both feet together, ensuring that the sole of the left foot touches the ground while the toes of the right foot touch the ground.</li>
		<li>Have subjects repeat this sequence until the end of the ladder.</li>
	</ol>
	</li>
	<li>Sequence 3A: Have participants perform the drill with diagonal and lateral movements.
	<ol>
		<li>Have them place their left foot, followed by the right foot, outside the first square to the left.</li>
		<li>Then, have them put the right foot in the first square, followed by the left foot. They will then place the right foot outside the second square to the right, followed by the left foot.</li>
		<li>Have subjects continue this sequence until the end of the ladder (Figure 7).</li>
	</ol>
	</li>
	<li>Sequence 3B: Use this sequence to increase the difficulty level depending on individual adaptation to sequence 3A.
	<ol>
		<li>Have participants position their left foot outside the ladder next to the first square to the left, and then the right foot in the first square, followed by the left foot, maintaining both feet together. Then, they will put the right foot outside the second square to the right and then the left foot in the second square, followed by the right foot, keeping both feet together (Figure 8).</li>
	</ol>
	</li>
	<li>Sequence 4: Have participants place their right foot in the first square, the left foot outside the ladder to the left next to the rung separating the first and second squares, and the right foot outside the ladder across from the left foot.
	<ol>
		<li>Have participants put the left foot in the second square, the right foot outside the ladder to the right next to the rung separating the second and third squares, and the left foot outside the ladder opposite the right foot. Have the subjects follow this pattern until the end of the ladder (Figure 9).</li>
	</ol>
	</li>
</ol>

<ol>
	<li>&#193;lvarez-Garc&#237;a, J., Dur&#225;n-S&#225;nchez, A., Del R&#237;o-Rama, M. C., Garc&#237;a-V&#233;lez, D. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Active+ageing:+Mapping+of+scientific+coverage.">Active ageing: Mapping of scientific coverage.</a> International Journal of Environmental Research and Public Health. 15 (12), E2727 (2018).</li><li>Weil, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Aging+And+Health.">Aging And Health.</a> Health Affairs. 34 (1), 6 (2018).</li><li>Sheppard, J., Young, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Agility+literature+review:+Classifications,+training+and+testing.">Agility literature review: Classifications, training and testing.</a> Journal of Sports Sciences. 24 (9), 919-932 (2006).</li><li>Paul, D. J., Gabbett, T. J., Nassis, G. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Agility+in+Team+Sports:+Testing,+Training+and+Factors+Affecting+Performance.">Agility in Team Sports: Testing, Training and Factors Affecting Performance.</a> Sports Medicine. 46 (3), 421-442 (2016).</li><li>Okubo, Y., Schoene, D., Lord, S. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Step+training+improves+reaction+time,+gait+and+balance+and+reduces+falls+in+older+people:+A+systematic+review+and+meta-analysis.">Step training improves reaction time, gait and balance and reduces falls in older people: A systematic review and meta-analysis.</a> British Journal of Sports Medicine. 51 (7), 586-593 (2017).</li><li>Liu-Ambrose, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Resistance+and+Agility+Training+Reduce+Fall+Risk+in+Women+Aged+75+to+85+with+Low+Bone+Mass:+A+6-Month+Randomized,+Controlled+Trial.">Resistance and Agility Training Reduce Fall Risk in Women Aged 75 to 85 with Low Bone Mass: A 6-Month Randomized, Controlled Trial.</a> Journal of the American Geriatrics Society. 52 (5), 657-665 (2004).</li><li>Liu-Ambrose, T., Khan, K. M., Eng, J. J., Lord, S. R., McKay, H. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Balance+confidence+improves+with+resistance+or+agility+training:+Increase+is+not+correlated+with+objective+changes+in+fall+risk+and+physical+abilities.">Balance confidence improves with resistance or agility training: Increase is not correlated with objective changes in fall risk and physical abilities.</a> Gerontology. 50 (6), 373-382 (2004).</li><li>Donath, L., van Die&#235;n, J., Faude, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Exercise-Based+Fall+Prevention+in+the+Elderly:+What+About+Agility?.">Exercise-Based Fall Prevention in the Elderly: What About Agility?.</a> Sports Medicine. 46 (2), 143-149 (2016).</li><li>Diamond, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Executive+Functions.">Executive Functions.</a> Annual Review of Clinical Psychology. 64 (1), 135-168 (2014).</li><li>Shigematsu, R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Square-Stepping+Exercise+and+Fall+Risk+Factors+in+Older+Adults:+A+Single-Blind,+Randomized+Controlled+Trial.">Square-Stepping Exercise and Fall Risk Factors in Older Adults: A Single-Blind, Randomized Controlled Trial.</a> Journals of Gerontology Series A Biological Sciences &amp; Medical Sciences. 63 (1), 76-82 (2008).</li><li>Teixeira, C. V. L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+square-stepping+exercise+and+basic+exercises+on+functional+fitness+of+older+adults.">Effect of square-stepping exercise and basic exercises on functional fitness of older adults.</a> Geriatrics and Gerontology International. 13 (4), 842-848 (2013).</li><li>Ng, R., Cheung, C. W., Kim Wai Sum, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+6-week+agility+ladder+drills+during+recess+intervention+on+dynamic+balance+performance.">Effects of 6-week agility ladder drills during recess intervention on dynamic balance performance.</a> Journal of Physical Education and Sport. 17 (1), 306-311 (2017).</li><li>Pawar, S. B., Pradeep, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+ladder+drills+training+in+female+kabaddi+players.">Effect of ladder drills training in female kabaddi players.</a> International Journal of Physical Education, Sports and Health. 180 (2), 180-184 (2018).</li><li>Raya, M. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparison+of+three+agility+tests+with+male+servicemembers:+Edgren+Side+Step+Test,+T-Test,+and+Illinois+Agility+Test.">Comparison of three agility tests with male servicemembers: Edgren Side Step Test, T-Test, and Illinois Agility Test.</a> Journal of Rehabilitation Research and Development. 50 (7), 951-960 (2013).</li><li>Buatois, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Five+times+sit+to+stand+test+is+a+predictor+of+recurrent+falls+in+healthy+community-living+subjects+aged+65+and+older.">Five times sit to stand test is a predictor of recurrent falls in healthy community-living subjects aged 65 and older.</a> Journal of the American Geriatrics Society. 56 (8), 1575-1577 (2008).</li><li>Podsiadlo, D., Richardson, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Timed+"Up+&amp;+Go":+A+Test+of+Basic+Functional+Mobility+for+Frail+Elderly+Persons.">The Timed "Up &amp; Go": A Test of Basic Functional Mobility for Frail Elderly Persons.</a> Journal of the American Geriatrics Society. 39 (2), 142-148 (1991).</li><li>Yamada, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Prevalence+of+sarcopenia+in+community-dwelling+Japanese+older+adults.">Prevalence of sarcopenia in community-dwelling Japanese older adults.</a> Journal of the American Medical Directors Association. 14 (12), 911-915 (2013).</li><li>Briggs, R. C., Gossman, M. R., Birch, R., Drews, J. E., Shaddeau, S. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Balance+performance+among+noninstitutionalized+elderly+women.">Balance performance among noninstitutionalized elderly women.</a> Physical Therapy. 69 (9), 748-756 (1989).</li><li>Yamada, M., Tanaka, B., Nagai, K., Aoyama, T., Ichihashi, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rhythmic+stepping+exercise+under+cognitive+conditions+improves+fall+risk+factors+in+community-dwelling+older+adults:+Preliminary+results+of+a+cluster-randomized+controlled+trial.">Rhythmic stepping exercise under cognitive conditions improves fall risk factors in community-dwelling older adults: Preliminary results of a cluster-randomized controlled trial.</a> Aging &amp; Mental Health. 15 (5), 647-653 (2011).</li><li>Reid, K. F., Fielding, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Skeletal+muscle+power:+A+critical+determinant+of+physical+functioning+in+older+adults.">Skeletal muscle power: A critical determinant of physical functioning in older adults.</a> Exercise and Sport Sciences Reviews. 40 (1), 4-12 (2012).</li><li>Bouaziz, W., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Health+benefits+of+multicomponent+training+programmes+in+seniors:+a+systematic+review.">Health benefits of multicomponent training programmes in seniors: a systematic review.</a> International Journal of Clinical Practice. 70 (7), 520-536 (2016).</li></ol>

The authors have nothing to disclose.

Several agility training programs are used for different populations, including sports teams4, children12, and older people6,10. These programs include square stepping exercises10,11 like the sequences described in the present study and can significantly improve the overall physical fitness of older adults by decreasing the risk of falls, fractures, immobili...

Being active while aging can optimize opportunities to improve physical, social, and mental health, enabling older people to participate actively in society1. Older adults should be payed attention to because of the consequences of aging2. People worldwide are living longer, and life expectancy is greater than 70 years for the first time in history. The number of individuals aged 60 years or older is increasing rapidly, and the health of these populations is usually poor. Aging impairs physical functions, mental capacity, and social interactions as a direct result of diseases or syndromes2.
Decrease in physical function may lead to falls, fractures, less access to health care, depression, and poor quality of life. Agility is crucial to prevent falls and maintain independence in older people. Sheppard and Young3 reported that agility involved &#34;whole-body movements in different directions in response to a stimulus.&#34;
Most studies evaluate agility in high-performance athletes4, although a few studies assess this parameter in older adults. Agility training for older people comprises preplanned gait adaptability training and reaction training (i.e., response to a signal)5. Liu-Ambrose et al.6,7 found that agility training improved postural stability, balance, and balance confidence, and decreased the risk of falls in older people. Moreover, this type of training is integrative and improves cognitive functions (e.g., perception, mental flexibility, and working memory) as well as neuromuscular and cardiovascular functions8,9.
Agility training is characterized by a set of exercise drills and can be automated. However, a physical trainer/instructor is necessary to create and teach the drills and indicate the right moment to increase the level of difficulty. Some studies found that step training improved physical fitness (e.g., balance, agility, lower limb strength and power, and decreased risk of falls) and cognitive function (e.g., choice stepping reaction time, inhibitory action, and executive function)5,10,11. The present study uses an agility ladder as a cost-effective instrument to assess the effects of preplanned gait training on the physical fitness of independent community-dwelling older adults.
The agility ladder used in the present study was previously employed to train school children and athletes12,13. The ladder had a length of 4.8 meters and 12 transverse rungs and was drawn on the floor using a 2 cm wide adhesive tape (Figure 1). Building a ladder on the floor avoided interruptions during training12 and decreased the risk of injuries. The study population was healthy, with a mean age of 66.9 &#177; 5.0 years, and lived independently in a community setting. The participants received 30 minute training sessions twice a week for 14 weeks.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Bluetooth speaker</td>
			<td>JBL</td>
			<td>-</td>
			<td>Used to time all protocol and signal the 30 s of each subset</td>
		</tr>
		<tr>
			<td>Chair</td>
			<td>-</td>
			<td>-</td>
			<td>Used on assessments</td>
		</tr>
		<tr>
			<td>Clipboard</td>
			<td>Tilibra</td>
			<td>-</td>
			<td>Used to support documents</td>
		</tr>
		<tr>
			<td>Crepe Tape of 2cm</td>
			<td>Dexter</td>
			<td>-</td>
			<td>Used to create the agility ladder on the floor</td>
		</tr>
		<tr>
			<td>Measuring Tape</td>
			<td>Stakley</td>
			<td>-</td>
			<td>Used to measure assessments distance</td>
		</tr>
		<tr>
			<td>Pen</td>
			<td>Bic</td>
			<td>-</td>
			<td>Used to note data</td>
		</tr>
		<tr>
			<td>Saucer Cone</td>
			<td>Kipsta</td>
			<td>-</td>
			<td>Used on assessments</td>
		</tr>
		<tr>
			<td>Stowatch</td>
			<td>Vollo</td>
			<td>-</td>
			<td>Used to time assessments results</td>
		</tr>
		<tr>
			<td>Sulfite Paper</td>
			<td>Chamex</td>
			<td>-</td>
			<td>used to write all data</td>
		</tr>
	</tbody>
</table>

a training program using an agility ladder for community-dwelling older adults

Aging impairs physical and cognitive functions and limits daily activities. Agility training can improve or maintain physical functioning in older people. The purpose of this study is to report the physical fitness benefits of a training program for independent community-dwelling older adults using an agility ladder. Each training session lasted approximately 30 minutes, and the benefits were achieved with two sessions per week for 14 weeks. Training was timed and involved four different drills and varying levels of difficulty through time. The exercises were performed at the School of Physical Education of the University of Campinas, S&#227;o Paulo state, Brazil. The study participants (n = 16; mean age of 66.9 &#177; 5.0 years) were instructed to perform the exercises as quickly as possible without making mistakes and were assisted by a physical trainer when they made mistakes. Assessments were performed both before and after training using five functional tests (i.e., Illinois agility, five times sit-to-stand, timed up-and-go, walking usual speed, and one-leg stand). Although the study sample was not compared with a control group, the results indicate that training protocols using an agility ladder are easy and practical and improve physical function performance in older adults.

Sixteen male and female independent community-living older people with a mean age of 66.9 &#177; 5.0 years were enrolled in the study. The interventions and assessments were performed at the School of Physical Education of UNICAMP.
Statistical analysis was performed using Minitab 18 software. The Mann-Whitney test was used to compare the functional test results before and after training. P values smaller than 0.05 were considere...

Watch this Scientific Journal Video about A Training Program Using an Agility Ladder for Community-Dwelling Older Adults at JoVE.com

A Training Program Using an Agility Ladder for Community-Dwelling Older Adults

The aim of this study is to present an agility training program for older people. The feasibility of this program is described, and the training protocol demonstrated by video imaging.

Aging impairs physical and cognitive functions and limits daily activities. Agility training can improve or maintain physical functioning in older people. ...

a-training-program-using-an-agility-ladder-for-community-dwelling

Research

JoVE Journal

Medicine

10.2K Views.  University of Campinas, UNICAMP. The aim of this study is to present an agility training program for older people. The feasibility of this program is described, and the training protocol demonstrated by video imaging.

Video: A Training Program Using an Agility Ladder for Community-Dwelling Older Adults

A Murine Model of Muscle Training by Neuromuscular Electrical Stimulation

Neuromuscular electrical stimulation (NMES) is a common clinical modality that is widely used to restore1, maintain2 or enhance3-5 muscle functional capacity. Transcutaneous surface stimulation of skeletal muscle involves a current flow between a cathode and an anode, thereby inducing excitement of the motor unit and the surrounding muscle fibers. 
NMES is an attractive modality to evaluate skeletal muscle adaptive responses for several reasons. First, it provides a reproducible experimental model in which physiological adaptations, such as myofiber hypertophy and muscle strengthening6, angiogenesis7-9, growth factor secretion9-11, and muscle precursor cell activation12 are well documented. Such physiological responses may be carefully titrated using different parameters of stimulation (for Cochrane review, see 13). In addition, NMES recruits motor units non-selectively, and in a spatially fixed and temporally synchronous manner14, offering the advantage of exerting a treatment effect on all fibers, regardless of fiber type. Although there are specified contraindications to NMES in clinical populations, including peripheral venous disorders or malignancy, for example, NMES is safe and feasible, even for those who are ill and/or bedridden and for populations in which rigorous exercise may be challenging. 
Here, we demonstrate the protocol for adapting commercially available electrodes and performing a NMES protocol using a murine model. This animal model has the advantage of utilizing a clinically available device and providing instant feedback regarding positioning of the electrode to elicit the desired muscle contractile effect. For the purpose of this manuscript, we will describe the protocol for muscle stimulation of the anterior compartment muscles of a mouse hindlimb.

A murine model of neuromuscular electrical stimulation (NMES), a safe and inexpensive clinical modality, to the anterior compartment muscles is described. This model has the advantage of modifying a readily available clinical device for the purpose of eliciting targeted and specific muscle contractions in mice.

Neuromuscular electrical stimulation (NMES) is a common clinical modality that is widely used to restore1, maintain2 or enhance3-5 muscle functional ...

Using Quantitative Real-time PCR to Determine Donor Cell Engraftment in a Competitive Murine Bone Marrow Transplantation Model

Murine bone marrow transplantation models provide an important tool in measuring hematopoietic stem cell (HSC) functions and determining genes/molecules that regulate HSCs. In these transplant model systems, the function of HSCs is determined by the ability of these cells to engraft and reconstitute lethally irradiated recipient mice. Commonly, the donor cell contribution/engraftment is measured by antibodies to donor- specific cell surface proteins using flow cytometry. However, this method heavily depends on the specificity and the ability of the cell surface marker to differentiate donor-derived cells from recipient-originated cells, which may not be available for all mouse strains. Considering the various backgrounds of genetically modified mouse strains in the market, this cell surface/ flow cytometry-based method has significant limitations especially in mouse strains that lack well-defined surface markers to separate donor cells from congenic recipient cells. Here, we reported a PCR-based technique to determine donor cell engraftment/contribution in transplant recipient mice. We transplanted male donor bone marrow HSCs to lethally irradiated congenic female mice. Peripheral blood samples were collected at different time points post transplantation. Bone marrow samples were obtained at the end of the experiments. Genomic DNA was isolated and the Y chromosome specific gene, Zfy1, was amplified using quantitative Real time PCR. The engraftment of male donor-derived cells in the female recipient mice was calculated against standard curve with known percentage of male vs. female DNAs. Bcl2 was used as a reference gene to normalize the total DNA amount. Our data suggested that this approach reliably determines donor cell engraftment and provides a useful, yet simple method in measuring hematopoietic cell reconstitution in murine bone marrow transplantation models. Our method can be routinely performed in most laboratories because no costly equipment such as flow cytometry is required.

Determining donor cell engraftment presents a challenge in mouse bone marrow transplant models that lack well-defined phenotypical markers. We described a methodology to quantify male donor cell engraftment in female transplant recipient mice. This method can be used in all mouse strains for the study of HSC functions.

Murine bone marrow transplantation models provide an important tool in measuring hematopoietic stem cell (HSC) functions and determining genes/molecules ...

Renal Capsule Xenografting and Subcutaneous Pellet Implantation for the Evaluation of Prostate Carcinogenesis and Benign Prostatic Hyperplasia

New therapies for two common prostate diseases, prostate cancer (PrCa) and benign prostatic hyperplasia (BPH), depend critically on experiments evaluating their hormonal regulation. Sex steroid hormones (notably androgens and estrogens) are important in PrCa and BPH; we probe their respective roles in inducing prostate growth and carcinogenesis in mice with experiments using compressed hormone pellets. Hormone and/or drug pellets are easily manufactured with a pellet press, and surgically implanted into the subcutaneous tissue of the male mouse host. We also describe a protocol for the evaluation of hormonal carcinogenesis by combining subcutaneous hormone pellet implantation with xenografting of prostate cell recombinants under the renal capsule of immunocompromised mice. Moreover, subcutaneous hormone pellet implantation, in combination with renal capsule xenografting of BPH tissue, is useful to better understand hormonal regulation of benign prostate growth, and to test new therapies targeting sex steroid hormone pathways.

We describe the manufacture of compressed hormone pellets, as well as subcutaneous surgical implantation into mice. This strategy can be combined with the growth of cell and tissue xenografts under the renal capsule of athymic nude mice to evaluate hormonal carcinogenesis and regulation of benign prostate growth.

New therapies for two common prostate diseases,  prostate cancer (PrCa) and benign prostatic hyperplasia (BPH), depend  critically on experiments ...

Collecting Saliva and Measuring Salivary Cortisol and Alpha-amylase in Frail Community Residing Older Adults via Family Caregivers

Salivary measures have emerged in bio-behavioral research that are easy-to-collect, minimally invasive, and relatively inexpensive biologic markers of stress. This article we present the steps for collection and analysis of two salivary assays in research with frail, community residing older adults-salivary cortisol and salivary alpha amylase. The field of salivary bioscience is rapidly advancing and the purpose of this presentation is to provide an update on the developments for investigators interested in integrating these measures into research on aging. Strategies are presented for instructing family caregivers in collecting saliva in the home, and for conducting laboratory analyses of salivary analytes that have demonstrated feasibility, high compliance, and yield quality specimens. The protocol for sample collection includes: (1) consistent use of collection materials; (2) standardized methods that promote adherence and minimize subject burden; and (3) procedures for controlling certain confounding agents. We also provide strategies for laboratory analyses include: (1) saliva handling and processing; (2) salivary cortisol and salivary alpha amylase assay procedures; and (3) analytic considerations.

We demonstrate: (1) procedures for collection of salivary samples in cognitive impaired older adults by family caregivers in the home setting, (2) procedures for measuring stress activity via salivary cortisol and alpha amylase, and (3) representative profiles. Protocols that allow researchers to study stress-linked processes advance our understanding of biological sensitivity and susceptibility.

Salivary measures have emerged in bio-behavioral research  that are easy-to-collect, minimally invasive, and relatively inexpensive  biologic markers of ...

Clinical Assessment of Spatiotemporal Gait Parameters in Patients and Older Adults

Spatial and temporal characteristics of human walking are frequently evaluated to identify possible gait impairments, mainly in orthopedic and neurological patients1-4, but also in healthy older adults5,6. The quantitative gait analysis described in this protocol is performed with a recently-introduced photoelectric system (see&#160;Materials table) which has the potential to be used in the clinic because it is portable, easy to set up (no subject preparation is required before a test), and does not require maintenance and sensor calibration. The photoelectric system consists of series of high-density floor-based photoelectric cells with light-emitting and light-receiving diodes that are placed parallel to each other to create a corridor, and are oriented perpendicular to the line of progression7. The system simply detects interruptions in light signal, for instance due to the presence of feet within the recording area. Temporal gait parameters and 1D spatial coordinates of consecutive steps are subsequently calculated to provide common gait parameters such as step length, single limb support and walking velocity8, whose validity against a criterion instrument has recently been demonstrated7,9. The measurement procedures are very straightforward; a single patient can be tested in less than 5 min&#160;and a comprehensive report can be generated in less than 1&#160;min.

This protocol is used to evaluate spatial and temporal gait variables of neurological/orthopedic patients and older persons by means of a recently-introduced floor-based photocell system.

Spatial and temporal characteristics of human walking are frequently evaluated to identify possible gait impairments, mainly in orthopedic and ...

Establishment of a Clinic-based Biorepository

The incidence of skin cancer (e.g., squamous cell carcinoma, basal cell carcinoma, and melanoma) has been increasing over the past several years. It is expected that there will be a parallel demand for cutaneous tumor samples for biomedical research studies. Tissue availability, however, is limited due the cost of establishing a biorepository and the lack of protocols available for obtaining clinical samples that do not interfere with clinical operations. A protocol was established to collect and process cutaneous tumor and associated blood and saliva samples that has minimal impact on routine clinical procedures on the date of a Mohs surgery. Tumor samples are collected and processed from patients undergoing their first layer of Mohs surgery for biopsy-proven cutaneous malignancies by the Mohs histotechnologist. Adjacent normal tissue is collected at the time of surgical closure. Additional samples that may be collected are whole-blood and buccal swabs. By utilizing tissue samples that are normally discarded, a biorepository was generated that offers several key advantages by being based in the clinic versus the laboratory setting. These include a wide range of collected samples; access to de-identified patient records, including pathology reports; and, for the typical donor, access to additional samples during follow-up visits.

Cutaneous tumors are often discarded following Mohs micrographic surgery. A protocol is described here that enables clinical support staff to effectively process and store cutaneous tumor (e.g., squamous cell carcinoma, basal cell carcinoma, and melanoma) samples for downstream laboratory applications without interfering with clinical operations.

The incidence of skin cancer (e.g., squamous cell carcinoma, basal cell carcinoma, and melanoma) has been increasing over the past several years. It is ...

Multi-Modal Home Sleep Monitoring in Older Adults

The gold standard for sleep monitoring is attended in-lab polysomnography; however, this method may be cost-prohibitive and inconvenient for patients and research participants. Home sleep testing has gained momentum in the field of sleep medicine due to its convenience and lower cost, as well as being more naturalistic. The accuracy and quality of home sleep testing, however, may be variable because studies are not monitored by sleep technologists. There has been some success in improving the accuracy of home sleep studies by having trained sleep technicians assist participants inside their homes with putting on the devices, but this can be intrusive and time-consuming for those involved. In this protocol, participants undergo at-home sleep monitoring with multiple devices: 1) a single-channel EEG device; 2) a home sleep test for sleep-disordered breathing and periodic limb movements; 3) actigraphy; and 4) sleep logs. A major challenge of this study is obtaining high-quality sleep monitoring data on the first attempt in order to minimize participant burden. This protocol describes the implementation of educational manuals with step-by-step instructions and photos. The goal is to improve the quality of home sleep testing.

Here, we present a protocol to improve the quality of data from home sleep testing by providing a method to enhance instructions through a structured participant visit. This protocol includes the implementation of a step-by-step educational manual with photos to ensure proper placement of equipment.

The gold standard for sleep monitoring is attended in-lab polysomnography; however, this method may be cost-prohibitive and inconvenient for patients and ...

A Computerized Functional Skills Assessment and Training Program Targeting Technology Based Everyday Functional Skills

Today, many functional skills are technology-based, so development of a technology-based training program has broad importance. Here we present a computerized functional skills training program that was paired in half of the participants with a commercially available cognitive training (CCT) program.
Non-impaired older individuals (NC) aged 60+ (n=45) and similarly aged individuals with mild cognitive impairment (MCI; n=50) were randomized to receive 12 weeks of twice-weekly computerized functional skills training (CFST) or 12 weeks of twice-weekly sessions split between CCT and CFST. Skills trained were use of an ATM; internet banking; ticket kiosk; telephone and internet prescription refill; medication management; and internet shopping. As with previous functional capacity assessments, we focus on completion time for each simulation.
51 participants completed the training program, either by mastering all 6 tasks (34) or completing 12 weeks of training. 44 more participants completed 4 or more training sessions so they were also analyzed for improvement up to their last training session. Completion time for all 6 tests significantly improved from the baseline assessment to the final training session in both groups of participants (all p&#60;0.001 with an average improvement in task completion time of 45%). Further, there was no differential improvement in MCI and NC in the 6 tests from baseline to end of training (all t&#60;1.66, all p&#62;0.12). Finally, combined CCT plus CFST did not differ from CSFT alone on any of the percent-change score measures (all t&#60;1.64, all p&#62;0.11).
Both NC and MCI groups evidenced substantial improvements in performance. CCT supplementation led to similar functional gains with half as many training sessions. The NC participants proceeded through the training fairly rapidly even without CCT supplementation; MCI participants required more training but learned equivalently. These findings suggest that even in cases with memory impairments, functional skills can be efficiently learned with training.

This training protocol uses computerized training to teach technology-related everyday functional skills. These skills include financial skills, travel and transit, as well as medication management.

Today, many functional skills are technology-based, so development of a technology-based training program has broad importance. Here we present a ...

Oral Health Assessment by Lay Personnel for Older Adults

Oral health is an often-undervalued contributor to overall health. The literature, however, underscores the myriad of systemic diseases influenced by oral health, including type II diabetes, heart disease, and atherosclerosis. Thus, assessments of oral health, called oral screenings, have a significant role in assessing risk of disease, managing disease, and even improving disease by oral care. Here we present a method to assess oral health quickly and consistently across time. The protocol is simple enough for non-oral health professionals such as students, family, and caregivers. Useful for any age of patient, the method is particularly key for older individuals who are often at risk of inflammation and chronic disease. Components of the method include existing oral health assessment scales and inventories, which are combined to produce a comprehensive assessment of oral health. Thus, oral characteristics assessed include intraoral and extraoral structures, soft and hard tissues, natural and artificial teeth, plaque, oral functions such as swallowing, and the impact this oral health status has on the patient&#39;s quality of life. Advantages of this method include its inclusion of measures and perceptions of both the observer and patient, and its ability to track changes in oral health over time. Results acquired are quantitative totals of questionnaire and oral screening items, which can be summed for an oral health status score. The scores of successive oral screenings can be used to track the progression of oral health across time and guide recommendations for both oral and overall health care.

The goal of this protocol is to enable non-dental professionals to assess oral health status for research or health-screening purposes. Aspects assessed include lips, tongue, soft and hard tissues, natural and artificial teeth, oral cleanliness, plaque, swallowing, and impact of oral health on quality of life.

Oral health is an often-undervalued contributor to overall health. The literature, however, underscores the myriad of systemic diseases influenced by oral ...

Development of a Novel Task-oriented Rehabilitation Program using a Bimanual Exoskeleton Robotic Hand

A robot-assisted hand is used for the rehabilitation of patients with impaired upper limb function, particularly for stroke patients with a loss of motor control. However, it is unclear how conventional occupational training strategies can be applied to the use of rehabilitation robots. Novel robotic technologies and occupational therapy concepts are used to develop a protocol that allows patients with impaired upper limb function to grasp objects using their affected hand through a variety of pinching and grasping functions. To conduct this appropriately, we used five types of objects: a peg, a rectangular cube, a cube, a ball, and a cylindrical bar. We also equipped the patients with a robotic hand, the Mirror Hand, an exoskeleton&#160;hand that is fitted to the subject&#8217;s affected hand and follows the movement of the sensor glove fitted to their unaffected hand (bimanual movement training (BMT)). This study had two stages. Three healthy subjects were first recruited to test the feasibility and acceptability of the training program. Three patients with hand dysfunction caused by stroke were then recruited to confirm the feasibility and acceptability of the training program, which was conducted on 3 consecutive days. On each day, the patient was monitored during 5 min of movement in a passive range of motion, 5 min of robot-assisted bimanual movement, and task-oriented training using the five objects. The results showed that both healthy subjects and subjects who had suffered a stroke in conjunction with the robotic hand could successfully grasp the objects. Both healthy subjects and those who had suffered a stroke performed well with the robot-assisted task-oriented training program in terms of feasibility and acceptability.

This study reports the development of a novel robot-assisted task-oriented program for hand rehabilitation. The developmental process consists of experiments using both healthy subjects and subjects who have had a stroke and suffered from subsequent motor control dysfunction.

A robot-assisted hand is used for the rehabilitation of patients with impaired upper limb function, particularly for stroke patients with a loss of motor ...