Name: lower limb biomechanical analysis of healthy participants
Uploaded: 2020-04-15T14:00:00
Description: Watch this Scientific Journal Video about Lower Limb Biomechanical Analysis of Healthy Participants at JoVE.com

We would like to thank Dr. Johnathan Williams for his advice on MATLAB data processing.

The method reported was followed in a study that received ethical approval from the Bournemouth University Research Ethics Committee (Reference 15005).
1. Participants
<ol>
	<li>Recruit healthy adults (aged from 23 to 63 years, mean &#177; S.D.; 42.0 &#177; 13.4, body mass 70.4 &#177; 15.3 kg, height 175.5 &#177; 9.8 cm; 15 males, 15 females) to participate in the study. Thirty participants were recruited for this study.</li>
	<li>Ensure that there is no self-reported history of dizziness, b...

<ol>
	<li>Lu, T. W., Chang, C. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biomechanics+of+human+movement+and+its+clinical+applications.">Biomechanics of human movement and its clinical applications.</a> The Kaohsiung Journal of Medical Sciences. 28 (2 Suppl), S13-S25 (2012).</li><li>Kaufman, K., An, K., Firestein, G. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Kelley and Firestein's Textbook of Rheumatology (Tenth Edition). , 78-89 (2017).</li><li>Sloot, L. H., van der Krogt, M. M., Harlaar, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Self-paced+versus+fixed+speed+treadmill+walking.">Self-paced versus fixed speed treadmill walking.</a> Gait &amp; Posture. 39 (1), 478-484 (2014).</li><li>Beaton, D. E., O'Driscoll, S. W., Richards, R. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Grip+strength+testing+using+the+BTE+work+simulator+and+the+jamar+dynamometer:+A+comparative+study.">Grip strength testing using the BTE work simulator and the jamar dynamometer: A comparative study.</a> The Journal of Hand Surgery. 20 (2), 293-298 (1995).</li><li>Jindal, P., Narayan, A., Ganesan, S., MacDermid, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Muscle+strength+differences+in+healthy+young+adults+with+and+without+generalized+joint+hypermobility:+a+cross-sectional+study.">Muscle strength differences in healthy young adults with and without generalized joint hypermobility: a cross-sectional study.</a> BMC Sports Science, Medicine &amp; Rehabilitation. 8, 12 (2016).</li><li>Muehlbauer, T., Granacher, U., Borde, R., Hortob&#225;gyi, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Non-Discriminant+Relationships+between+Leg+Muscle+Strength,+Mass+and+Gait+Performance+in+Healthy+Young+and+Old+Adults.">Non-Discriminant Relationships between Leg Muscle Strength, Mass and Gait Performance in Healthy Young and Old Adults.</a> Gerontology. 64 (1), 11-18 (2018).</li><li>van den Bogert, A. J., Geijtenbeek, T., Even-Zohar, O., Steenbrink, F., Hardin, E. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+real-time+system+for+biomechanical+analysis+of+human+movement+and+muscle+function.">A real-time system for biomechanical analysis of human movement and muscle function.</a> Medical &amp; Biological Engineering &amp; Computing. 51 (10), 1069-1077 (2013).</li><li>. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> HBM2 Reference Manual. , 9-11 (2017).</li><li>Sloot, L. H., van der Krogt, M. M., Harlaar, J. <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+adding+a+virtual+reality+environment+to+different+modes+of+treadmill+walking.">Effects of adding a virtual reality environment to different modes of treadmill walking.</a> Gait Posture. 39 (3), 939-945 (2014).</li><li>Liu, W. Y., 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=Reproducibility+and+Validity+of+the+6-Minute+Walk+Test+Using+the+Gait+Real-Time+Analysis+Interactive+Lab+in+Patients+with+COPD+and+Healthy+Elderly.">Reproducibility and Validity of the 6-Minute Walk Test Using the Gait Real-Time Analysis Interactive Lab in Patients with COPD and Healthy Elderly.</a> PLoS One. 11 (9), e0162444 (2016).</li><li>Herman, T., Mirelman, A., Giladi, N., Schweiger, A., Hausdorff, J. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Executive+Control+Deficits+as+a+Prodrome+to+Falls+in+Healthy+Older+Adults:+A+Prospective+Study+Linking+Thinking,+Walking,+and+Falling.">Executive Control Deficits as a Prodrome to Falls in Healthy Older Adults: A Prospective Study Linking Thinking, Walking, and Falling.</a> The Journals of Gerontology: Series A. 65 (10), 1086-1092 (2010).</li><li>Geijtenbeek, T., Steenbrink, F., Otten, B., Even-Zohar, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Proceedings+of+the+10th+International+Conference+on+Virtual+Reality+Continuum+and+Its+Applications+in+Industry.">Proceedings of the 10th International Conference on Virtual Reality Continuum and Its Applications in Industry.</a> , 201-208 (2011).</li><li>Zeni, J. A., Higginson, J. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gait+parameters+and+stride-to-stride+variability+during+familiarization+to+walking+on+a+split-belt+treadmill.">Gait parameters and stride-to-stride variability during familiarization to walking on a split-belt treadmill.</a> Clinical Biomechanics (Bristol, Avon). 25 (4), 383-386 (2010).</li><li>Meldrum, D., Cahalane, E., Conroy, R., Fitzgerald, D., Hardiman, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Maximum+voluntary+isometric+contraction:+reference+values+and+clinical+application.">Maximum voluntary isometric contraction: reference values and clinical application.</a> Amyotroph Lateral Sclerosis. 8 (1), 47-55 (2007).</li><li>Ancillao, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Modern Functional Evaluation Methods for Muscle Strength and Gait Analysis. , 133 (2018).</li><li>Mun, J. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+method+for+the+reduction+of+skin+marker+artifacts+during+walking+:+Application+to+the+knee.">A method for the reduction of skin marker artifacts during walking : Application to the knee.</a> KSME International Journal. 17 (6), 825-835 (2003).</li><li>Liu, P. C., Liu, J. F., Chen, L. Y., Xia, K., Wu, X. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intermittent+pneumatic+compression+devices+combined+with+anticoagulants+for+prevention+of+symptomatic+deep+vein+thrombosis+after+total+knee+arthroplasty:+a+pilot+study.">Intermittent pneumatic compression devices combined with anticoagulants for prevention of symptomatic deep vein thrombosis after total knee arthroplasty: a pilot study.</a> Therapeutics and Clinical Risk Management. 13, 179-183 (2017).</li><li>Al-Amri, M., Al Balushi, H., Mashabi, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intra-rater+repeatability+of+gait+parameters+in+healthy+adults+during+self-paced+treadmill-based+virtual+reality+walking.">Intra-rater repeatability of gait parameters in healthy adults during self-paced treadmill-based virtual reality walking.</a> Computer Methods in Biomechanics and Biomedical Engineering. 20 (16), 1669-1677 (2017).</li><li>Zeni, J., Richards, J., Higginson, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Two+simple+methods+for+determining+gait+events+during+treadmill+and+overground+walking+using+kinematic+data.">Two simple methods for determining gait events during treadmill and overground walking using kinematic data.</a> Gait &amp; Posture. 27 (4), 710-714 (2008).</li><li>Tsaopoulos, D. E., Baltzopoulos, V., Richards, P. J., Maganaris, C. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanical+correction+of+dynamometer+moment+for+the+effects+of+segment+motion+during+isometric+knee-extension+tests.">Mechanical correction of dynamometer moment for the effects of segment motion during isometric knee-extension tests.</a> Journal of Applied Physiology. 111 (1), 68-74 (2011).</li><li>Abernethy, P., Wilson, G., Logan, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Strength+and+power+assessment.+Issues,+controversies+and+challenges.">Strength and power assessment. Issues, controversies and challenges.</a> Sports Medicine. 19 (6), 401-417 (1995).</li><li>Kroll, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reliability+of+a+Selected+Measure+of+Human+Strength.">Reliability of a Selected Measure of Human Strength.</a> Research Quarterly, American Association for Health, Physical Education and Recreation. 33 (3), 410-417 (1962).</li><li>Anzak, A., Tan, H., Pogosyan, A., Brown, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Doing+better+than+your+best:+loud+auditory+stimulation+yields+improvements+in+maximal+voluntary+force.">Doing better than your best: loud auditory stimulation yields improvements in maximal voluntary force.</a> Experimental Brain Research. 208 (2), 237-243 (2011).</li><li>Belkhiria, C., De Marco, G., Driss, T. <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+verbal+encouragement+on+force+and+electromyographic+activations+during+exercise.">Effects of verbal encouragement on force and electromyographic activations during exercise.</a> Journal of Sports Medicine and Physical Fitness. 58 (5), 750-757 (2018).</li><li>Bickers, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Does+verbal+encouragement+work?+The+effect+of+verbal+encouragement+on+a+muscular+endurance+task.">Does verbal encouragement work? The effect of verbal encouragement on a muscular endurance task.</a> Clinical Rehabilitation. 7 (3), 196-200 (1993).</li><li>Karaba-Jakovljevic, D., Popadic-Gacesa, J., Grujic, N., Barak, O., Drapsin, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Motivation+and+motoric+tests+in+sports.">Motivation and motoric tests in sports.</a> Medicinki Pregled. 60 (5-6), 231-236 (2007).</li><li>Andreacci, J. 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=The+effects+of+frequency+of+encouragement+on+performance+during+maximal+exercise+testing.">The effects of frequency of encouragement on performance during maximal exercise testing.</a> Journal of Sports Science. 20 (4), 345-352 (2002).</li><li>Rendos, N. K., 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=Variations+in+Verbal+Encouragement+Modify+Isokinetic+Performance.">Variations in Verbal Encouragement Modify Isokinetic Performance.</a> Journal of Strength and Conditioning Research. 33 (3), 708-716 (2019).</li></ol>

The contribution of this study is to accurately and comprehensively describe within one protocol the techniques for combined gait analysis and muscle strength testing that have not previously been described together.
In order to achieve accurate results for gait analysis, there are two areas that require maximum attention: 1) marker placements and 2) acclimatization time. The accuracy of the measured data is heavily dependent on the accuracy of the model used. The other key factors that affect...

The discipline of biomechanics primarily involves the study of stress, strain, loads and motion of biological systems - solid and fluid alike. It also involves the modelling of mechanical effects on the structure, size, shape and movement of the body1. For many years, developments in this field have improved our understanding of normal and pathologic gait, mechanics of neuromuscular control, and mechanics of growth and form2.
The main objective of this article is to present a comprehensive methodology on two of the latest technologies available to measure lower limb biomechanics of individuals. The gait analysis system measures and quantifies gait biomechanics by using a self-paced (SP) treadmill in combination with an augmented reality environment, which integrates a SP algorithm to regulate the treadmill&#39;s speed, as described by Sloot et al3. The muscle strength testing equipment is used as an assessment and a treatment tool for upper extremity rehabilitation4. This device can objectively assess a variety of physiological patterns of movement or job simulation tasks in isometric and isotonic modes. It is currently recognized as the gold standard for upper limb strength measurement5 but the evidence related specifically to the lower limb remains unclear. This paper explains the detailed protocol for completing an assessment of gait and isometric strength for the lower extremity.
Within biomechanical analysis, it is useful to combine assessments of functional performance (such as gait analysis) with specific tests of muscular performance. This is because whilst it may be assumed that increased muscle strength improves functional performance, this may not always be apparent6. This understanding is required for the improved future design of rehabilitation protocols and research strategies to assess these approaches.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>701 Small lever</td>
			<td>Baltimore Therapeutic Equipment Company (BTE)</td>
			<td>Not Available - Online link provided in description</td>
			<td>The unique attachment designed for the Primus RS to measure Knee Extension/Flexion - <a href="https://store.btetech.com/collections/primus/products/701-small-lever" target="_blank">https://store.btetech.com/collections/primus/products/701-small-lever</a></td>
		</tr>
		<tr>
			<td>D-Flow Software - Vresion 3.26</td>
			<td>Motekforce Link</td>
			<td>Not Available - Online link provided in description</td>
			<td>Software used to control GRAIL system - <a href="https://summitmedsci.co.uk/products/motek-dflow-hbm-software/" target="_blank">https://summitmedsci.co.uk/products/motek-dflow-hbm-software/</a></td>
		</tr>
		<tr>
			<td>Gait Offline Analysis (GOAT) - Version 2.3</td>
			<td>Motekforce Link</td>
			<td>Not Available - Online link provided in description</td>
			<td>Software used for the analysis of the gait parameters - <a href="https://www.motekmedical.com/product/grail/" target="_blank">https://www.motekmedical.com/product/grail/</a></td>
		</tr>
		<tr>
			<td>Gait Real-time Analysis Interactive Lab (GRAIL)</td>
			<td>Motekforce Link</td>
			<td>Not Available - Online link provided in description</td>
			<td>GRAIL system measures and quantifies gait biomechanics by using a virtual reality based self-paced (SP) treadmill - <a href="https://www.motekmedical.com/product/grail/" target="_blank">https://www.motekmedical.com/product/grail/</a></td>
		</tr>
		<tr>
			<td>Leg Pad for 701</td>
			<td>Baltimore Therapeutic Equipment Company (BTE)</td>
			<td>Not Available - Online link provided in description</td>
			<td>The unique attachment designed for the Primus RS to measure Knee Extension/Flexion - <a href="https://store.btetech.com/collections/primus/products/701-802-leg-pad" target="_blank">https://store.btetech.com/collections/primus/products/701-802-leg-pad</a></td>
		</tr>
		<tr>
			<td>Positioning Chair</td>
			<td>Baltimore Therapeutic Equipment Company (BTE)</td>
			<td>Not Available - Online link provided in description</td>
			<td>Participant Positioning Chair is designed for assessment and treatment of the lower exteremeties. The chair is designed for multiple positions. <a href="https://www.btetech.com/product/primus/" target="_blank">https://www.btetech.com/product/primus/</a></td>
		</tr>
		<tr>
			<td>Primus RS</td>
			<td>Baltimore Therapeutic Equipment Company (BTE)</td>
			<td>Not Available - Online link provided in description</td>
			<td>Primus RS equipment captures and reports real time objective data in Isotonic, Isometric, and Isokinetic resistance modes - <a href="https://www.btetech.com/wp-content/uploads/BTE-Rehabilitation-Equipment-PrimusRS-Brochure-1.pdf" target="_blank">https://www.btetech.com/wp-content/uploads/BTE-Rehabilitation-Equipment-PrimusRS-Brochure-1.pdf</a></td>
		</tr>
	</tbody>
</table>

lower limb biomechanical analysis of healthy participants

Biomechanical analysis techniques are useful in the study of human movement. The aim of this study was to introduce a technique for the lower limb biomechanical assessment in healthy participants using commercially available systems. Separate protocols were introduced for the gait analysis and muscle strength testing systems. To ensure maximum accuracy for gait assessment, attention should be given to the marker placements and self-paced treadmill acclimatization time. Similarly, participant positioning, a practice trial, and verbal encouragement are three critical stages in muscle strength testing. The current evidence suggests that the methodology outlined in this article may be effective for the assessment of lower limb biomechanics.

The mean and standard deviation of the spatial-temporal, kinematics, and kinetic gait parameters are given in Table 2. MVIC data for all 30 participants are summarized in Table 3. A typical set of data for the left and right side of one participant showing graphical representation of gait parameters is provided in Figure 4 and Figure 5, respectively.
<p class="jove_content" fo:keep-together.w...

Watch this Scientific Journal Video about Lower Limb Biomechanical Analysis of Healthy Participants at JoVE.com

Lower Limb Biomechanical Analysis of Healthy Participants

This article introduces a comprehensive experimental methodology on two of the latest technologies available to measure the lower limb biomechanics of individuals.

Biomechanical analysis techniques are useful in the study of human movement. The aim of this study was to introduce a technique for the lower limb ...

Biomechanical analysis techniques are useful in the study of human movement. This technique can be used for leveling biomechanical assessment in healthy participants using commercially available systems. In biomechanical analysis, it's useful to have specific test of function like gait analysis combined with tests looking at muscular performance.Understanding the relationship between muscle strength and the ability to perform functional tasks such as walking is needed in the future design of research studies and rehabilitation protocols. Demonstrating our protocol today is Francesco Ferraro, trial manager in our team. Before beginning the analysis, confirm that the participant is wearing tight non-reflective clothing such as cycling shorts or leggings and use double-sided adhesive tape to attach 25 passive reflective markers onto the participant according to the lower body configuration of the human body model as illustrated.Use a joint ruler to take measurements of the required knee and ankle widths for the model and secure the participant to a safety harness that is fastened to an overhead frame. Start a new session in the database and make sure the session is active. Under the Subject tab, click the Labeling Skeleton button and navigate to the Lower Limb HBM N2 VST file to create a new participant.Enter the name of the participant and click OK.The name of the participant will appear in the subjects window. In the tools windows, open the Subject Preparation tab. Click to zero level the force plates and ask the participant to stand in the middle of the treadmill while walking at a comfortable speed for five minutes immediately following the acclimatization and click Start Recording in the software to begin recording the gait data.Stop the recording after acquiring the desired amount of data. To remove the high-frequency noise in the data, open the processing software, select a low-pass filter for the marker data, and determine the individual strides from the vertical force data using the foot markers to ascertain the gain events. Export the data as a CSV file.The gait parameters such as the kinematic, kinetic, and spatial temporal data can then be analyzed. To measure the participant's muscle strength based on the maximum voluntary isometric contraction, first attach the tool pad number 701 to the dynamometer exercise head and have the participant sit on the chair with a back rest. To test the knee isometric muscle strength, use the up/down switch to align the dynamometer axis with the anatomical axis of rotation of the knee and place the pad of the tool centrally at the lower part of the shin of the tibia.Have the participant position the knee at a 90 degree flexion with the hip in neutral rotation and abduction and the foot in plantar flexion. Then have the participant place their hands on their abdomen and stabilize the trunk, hips, and mid thigh on the chair with Velcro straps. To familiarize the participant with the testing maneuver, have the participant extend their knee before flexing to exert a maximum contraction on the command go for three seconds while providing verbal prompts and encouragement.When the participant is comfortable with the procedure, ensure the participant that they can stop the test immediately if they experience any unusual pain or discomfort and allow the participant for rest for two minutes. Then repeat the trial three times for the left leg and three times for the right leg as just demonstrated recording and exporting the data in Newtons. Here, the mean and standard deviation of the spatial temporal, kinematics, and kinetic gait parameters of 30 participants from a representative analysis are shown.As observed in this table summarizing the mean and standard deviation of the maximum voluntary isometric contraction for the knee joints of the participants, on average, the right knee demonstrated a slightly more robust extension and flexion in the assessed participants. In this offline analysis of the gait assessment of one participant, the spatial temporal data and kinematic and kinetic gait cycle for the left side of the participant can be observed. These data were similarly reproduced when the spatial temporal data and kinematic and kinetic gait cycle for the right side of the participant were assessed.Clinical research has identified clear links between human biomechanics characteristics and different medical conditions. This protocol can be used for the accurate and comprehensive testing of combined gait analysis and muscle strength in a way that has not been previously described. Future research in orthopedic surgery will require accurate and repeatable techniques to measure gait.The data collected through the technique described is consistent with international reference data for gait analysis and isometric muscle strength testing.

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TRUE Gene Silencing: Screening of a Heptamer-type Small Guide RNA Library for Potential Cancer Therapeutic Agents

TRUE gene silencing (termed after tRNase ZL-utilizing efficacious gene silencing) is one of the RNA-directed gene silencing technologies, which utilizes an artificial small guide RNA (sgRNA) to guide tRNA 3&#8242; processing endoribonuclease, tRNase ZL, to recognize a target RNA. sgRNAs can be taken up by cells without any transfection reagents and can downregulate their target RNA levels and/or induce apoptosis in human cancer cells. We have screened an sgRNA library containing 156 heptamer-type sgRNAs for the effect on viability of human myeloma and leukemia cells, and found that 20 of them can efficiently induce apoptosis in at least one of the cancer cell lines. Here we present a protocol for screening of a heptamer-type sgRNA library for potential therapeutic drugs against blood cancers. The protocol includes how to construct the sgRNA library, how to assess the effect of each sgRNA on cell viability, and how to further evaluate the effective sgRNAs by flow cytometry. Around 2,000 hits would be expected to be obtained by screening the full-scale sgRNA library composed of 16,384 heptamers.

Here we present a protocol for screening of a heptamer-type sgRNA library for potential therapeutic drugs against blood cancers.

TRUE gene silencing (termed after tRNase ZL-utilizing efficacious gene silencing) is one of the RNA-directed gene silencing technologies, which utilizes ...

Oral Biofilm Sampling for Microbiome Analysis in Healthy Children

Oral biofilm and its molecular analysis provide a basis for investigating various dental research and clinical questions. Knowledge of biofilm composition leads to a better understanding of cariogenic and periopathogenic mechanisms. Microbial changes taking place in the oral cavity during childhood are of interest for several reasons. The evolution of the child oral microbiota and shifts in its composition need to be analyzed further to understand and possibly prevent the onset of disease. At the same time, advanced knowledge of the natural composition of oral biofilm is needed. Early stages of caries-free permanent dentition with healthy gums provide a widely unaffected subgingival habitat that can serve as an in situ baseline for studying features of oral health and disease. Analysis of children's oral biofilm during different stages in life is thus an important theme in the field. Modern molecular analysis methods can provide comprehensive information about the bacterial diversity of such biofilms. To enable microbiota data comparison, it is important to standardize each step in the procedure for molecular data generation. This procedure spans from clinical sampling, Next Generation Sequencing (NGS), bioinformatic data processing, to taxonomic interpretation. One of the most critical factors here is biofilm sampling. Sampling in children is even more challenging in particular due to limited space in subgingival areas. We thus focus on the use of paper points for subgingival sampling. This article provides a detailed protocol for oral biofilm sampling of the subgingival sulcus, the mucosa, and saliva in children.

Changes in the oral microbiome throughout childhood are of growing interest. Comparison of different microbiome studies reveals a lack of standardized sampling protocols. Limited space makes sampling the sound subgingival sulcus of children challenging. Paper point sampling is presented here in detail as the method of choice for this area.

Oral biofilm and its molecular analysis provide a basis for investigating various dental research and clinical questions. Knowledge of biofilm composition ...

Assessment of the Efficacy of An Osteopathic Treatment in Infants with Biomechanical Impairments to Suckling

Breastfeeding can be challenging for mother-infant dyads experiencing biomechanical suckling difficulties. Although lactation consultants (LCs) all over the world have increased their skills in this field and can provide support to help position the infant at the breast, the impact of their intervention might be limited in the presence of stiff structures in the infants. Here we present a protocol for a randomized controlled trial to assess the efficacy of osteopathic treatment, coupled with lactation consultation, for infants&#39; biomechanical suckling difficulties. It proposes a set-up and a sequence of actions to ensure an optimal context for treatment, as well as a blinding of parents and LCs to the intervention. Data such as the infant&#39;s latch ability measured with the LATCH Assessment Tool, the mother&#39;s nipple pain with a visual analog scale (VAS), and the mother&#39;s perceptions are collected by LCs four times over a 10-day period. Osteopathic lesions are documented by the osteopath, using a standardized assessment grid. Structures of interest are coherent with the anatomical zones involved in latching onto the breast. This protocol also proposes a strategy to document systematically an osteopathic profile of infants with biomechanical suckling difficulties in their first weeks of life. The implementation of this protocol confirms its feasibility for osteopathic assessment and treatment and paves the way for future trials to further explore the efficacy of osteopathic techniques for infants with biomechanical suckling difficulties.

Osteopathy is an emerging field of clinical research. Here we present a protocol to assess the efficacy of an osteopathic intervention coupled with lactation consultation, in infants with biomechanical issues impeding breastfeeding.

Breastfeeding can be challenging for mother-infant dyads experiencing biomechanical suckling difficulties. Although lactation consultants (LCs) all over ...

Preclinical Model of Hind Limb Ischemia in Diabetic Rabbits

Peripheral vascular disease is a widespread clinical problem that affects millions of patients worldwide. A major consequence of peripheral vascular disease is the development of ischemia. In severe cases, patients can develop critical limb ischemia in which they experience constant pain and an increased risk of limb amputation. Current therapies for peripheral ischemia include bypass surgery or percutaneous interventions such as angioplasty with stenting or atherectomy to restore blood flow. However, these treatments often fail to the continued progression of vascular disease or restenosis or are contraindicated due to the overall poor health of the patient. A promising potential approach to treat peripheral ischemia involves the induction of therapeutic neovascularization to allow the patient to develop collateral vasculature. This newly formed network alleviates peripheral ischemia by restoring perfusion to the affected area. The most frequently employed preclinical model for peripheral ischemia utilizes the creation of hind limb ischemia in healthy rabbits through femoral artery ligation. In the past, however, there has been a strong disconnect between the success of preclinical studies and the failure of clinical trials regarding treatments for peripheral ischemia. Healthy animals typically have robust vascular regeneration in response to surgically induced ischemia, in contrast to the reduced vascularity and regeneration in patients with chronic peripheral ischemia. Here, we describe an optimized animal model for peripheral ischemia in rabbits that includes hyperlipidemia and diabetes. This model has reduced collateral formation and blood pressure recovery in comparison to a model with a higher cholesterol diet. Thus, the model may provide better correlation with human patients with compromised angiogenesis from the common co-morbidities that accompany peripheral vascular disease.

We describe a surgical procedure used to induce peripheral ischemia in rabbits with hyperlipidemia and diabetes. This surgery acts as a preclinical model for conditions experienced in peripheral artery disease in patients. Angiography is also described as a means to measure the extent of introduced ischemia and recovery of perfusion.

Peripheral vascular disease is a widespread clinical problem that affects millions of patients worldwide. A major consequence of peripheral vascular ...

The Lower Body Positive Pressure Treadmill for Knee Osteoarthritis Rehabilitation

Here, based on a clinician&rsquo;s point-of-view, we propose a two-model lower body positive pressure (LBPP) protocol (walking and squatting models) in addition to a clinical, functional assessment methodology, including details for further encouragement of the development of non-drug surgical intervention strategies in knee osteoarthritis patients. However, we only present the effect of LBPP training in improvement of pain and knee function in one patient through three-dimensional gait analysis. The exact, long-term effects of this approach should be explored in future studies.

Here, based on a clinician&#8217;s point-of-view, we propose a two-model lower body positive pressure (LBPP) protocol (walking and squatting models) in addition to a clinical, functional assessment methodology, including details for further encouragement of the development of non-drug surgical intervention strategies in knee osteoarthritis patients. However, we only present the effect of LBPP training in improvement of pain and knee function in one patient through three-dimensional gait analysis. The exact, long-term effects of this approach should be explored in future studies.

Here, based on a clinician&rsquo;s point-of-view, we propose a two-model lower body positive pressure (LBPP) protocol (walking and squatting models) in ...

On-Site Sampling and Extraction of Brain Tumors for Metabolomics and Lipidomics Analysis

Despite the variety of tools available for cancer diagnosis and classification, methods that enable fast and simple characterization of tumors are still in need. In recent years, mass spectrometry has become a method of choice for untargeted profiling of discriminatory compound as potential biomarkers of a disease. Biofluids are generally considered as preferable matrices given their accessibility and easier sample processing while direct tissue profiling provides more selective information about a given cancer. Preparation of tissues for the analysis via traditional methods is much more complex and time-consuming, and, therefore, not suitable for fast on-site analysis. The current work presents a protocol combining sample preparation and extraction of small molecules on-site, immediately after tumor resection. The sampling device, which is of the size of an acupuncture needle, can be inserted directly into the tissue and then transported to the nearby laboratory for instrumental analysis. The results of metabolomics and lipidomics analyses demonstrate the capability of the approach for the establishment of phenotypes of tumors related to the histological origin of the tumor, malignancy, and genetic mutations, as well as for the selection of discriminating compounds or potential biomarkers. The non-destructive nature of the technique permits subsequent performance of routinely used tests e.g., histological tests, on the same samples used for SPME analysis, thus enabling attainment of more comprehensive information to support personalized diagnostics.

The manuscript presents on-site sampling of human brain tumors with solid phase microextraction followed by their biochemical profiling towards the discovery of biomarkers.

Despite the variety of tools available for cancer diagnosis and classification, methods that enable fast and simple characterization of tumors are still ...

Ultrasound Evaluation of Achilles Tendon Morphology

Measurement of Healthy and Injured Triceps Surae Morphology

Achilles tendon injuries occur throughout the lifespan and can negatively affect quality of life and overall health. Achilles tendinopathy is generally classified as an overuse injury associated with fusiform tendon thickening, neovascularization, and interstitial tendon degeneration. Current literature suggests these structural changes are associated with&#160;symptoms and lower physical activity levels, as well as&#160;symptoms and lower extremity function in the long term. Surgically and non-surgically managed Achilles tendon ruptures result in increased tendon cross-sectional area (CSA) and a lengthened Achilles tendon. Both structural outcomes have clinical implications, as larger CSA positively predicts function, whereas increased tendon lengthening predicts reduced function after Achilles tendon rupture. Given the relationship between structural changes associated with Achilles tendon injuries for both injury severity and injury recovery, it is critical to be able to quantify Achilles tendon structure reliably and accurately. Silbernagel&#39;s group has established a valid and reliable method for efficiently evaluating triceps surae muscle and tendon structure. In this protocol, B-mode musculoskeletal ultrasound imaging is used to measure triceps surae structure, including Achilles tendon thickness and CSA, soleus thickness, and the presence of additional findings (calcifications and bursitis). B-mode extended field-of-view is used to measure Achilles tendon length and gastrocnemius anatomical CSA. Finally, power Doppler is used to identify intratendinous neovascularization. Quantification of triceps surae structure allows for comparison between limbs as well as longitudinal changes in response to exercise and treatment for healthy individuals and those with Achilles tendon injuries. This protocol has been used in many research studies to date and proves valuable in understanding the relationship between tendon structure and injury development, severity, and recovery. As ultrasound devices are becoming more affordable and portable, this protocol proves promising as a clinical tool, given its quick and efficient methods.

Author Spotlight: Exploring the Complexities of Achilles Tendon Injuries &amp;#8212; Research and Future Directions

Ultrasound imaging is becoming more accessible in clinical and research settings, and a consistent protocol will be beneficial for comparison between studies and for clinical interpretations. This protocol for ultrasound evaluation is a valid and reliable method to evaluate Achilles tendon morphology in healthy, tendinopathic, and ruptured tendons.

Achilles tendon injuries occur throughout the lifespan and can negatively affect quality of life and overall health. Achilles tendinopathy is generally ...

Ultrasound in Evaluating Diaphragm Dysfunction and Monitoring Treatment

Measuring Diaphragm Thickness and Function Using Point-of-Care Ultrasound

The diaphragm is the main component of the respiratory muscle pump. Diaphragm dysfunction can cause dyspnea and exercise intolerance, and predisposes affected individuals to respiratory failure. In mechanically ventilated patients, the diaphragm is susceptible to atrophy and dysfunction through disuse and other mechanisms. This contributes to failure to wean and poor long-term clinical outcomes. Point-of-care ultrasound provides a valid and reproducible method for evaluating diaphragm thickness and contractile activity (thickening fraction during inspiration) that can be readily employed by clinicians and researchers alike. This article presents best practices for measuring diaphragm thickness and quantifying diaphragm thickening during tidal breathing or maximal inspiration. Once mastered, this technique can be used to diagnose and prognosticate diaphragm dysfunction, and guide and monitor response to treatment over time in both healthy individuals and acute or chronically ill patients.

Author Spotlight: Unraveling the Impact of Mechanical Ventilation on Diaphragm Function and Patient Outcomes 

Diaphragm thickness and function can be assessed in healthy individuals and critically ill patients using point-of-care ultrasound. This technique offers an accurate, reproducible, feasible, and well-tolerated method for evaluating diaphragm structure and function.

The diaphragm is the main component of the respiratory muscle pump. Diaphragm dysfunction can cause dyspnea and exercise intolerance, and predisposes ...

Rehabilitation Treatment Plan for ACLR Patients

Early Weight-Bearing Rehabilitation Protocol After Anterior Cruciate Ligament Reconstruction

Anterior cruciate ligament (ACL) injury is one of the common sports injuries. Anterior cruciate ligament reconstruction (ACLR) is the mainstream treatment for ACL injury, aiming to regain normal anatomical structure and stability of the knee joint and promote the patient's return to sports. Under the guidance of the concept of enhanced recovery after surgery, early weight-bearing rehabilitation (EWB) is an important factor affecting patient function and quality of life. However, there is no consensus on whether EWB rehabilitation can be performed after ACL surgery. 
This study aims to explore the safety and feasibility of EWB after ACL surgery. The study implemented a gradual EWB rehabilitation program in the experimental group, including weight-shifting training, balance training, and gait training on the affected lower limb, and assessed wound healing and stability of the knee joint. The study found that EWB after ACLR is safe and feasible. EWB rehabilitation not only does not pose a negative effect on the patient's knee pain, swelling, wound healing, and stability, but also helps to improve knee active flexion and quality of life faster and better. The EWB program in this study is simple, safe, and effective, and it provides strong theoretical guidance and practical demonstration for accelerated rehabilitation after ACLR.

Author Spotlight: Implementing the Enhanced Recovery After Surgery Concept in Rehabilitation Following Anterior Cruciate Ligament Reconstruction

Based on the safety and feasibility, this article presents an early weight-bearing rehabilitation protocol after anterior cruciate ligament reconstruction. The protocol is clear and easy to operate, which is helpful to promote its use in clinical practice and accelerate the functional recovery of patients.

Anterior cruciate ligament (ACL) injury is one of the common sports injuries. Anterior cruciate ligament reconstruction (ACLR) is the mainstream treatment ...

Evaluating Diaphragm Thickness and Thickening Fraction During Tidal Breathing in Healthy Participants

To begin, position the patient in a semi recumbent position on their back. Cover the tip of the linear array transducer with ultrasound gel. After ensuring the ultrasound is in B mode for positioning, hold the probe by enclosing the tip of the probe with the thumb and index finger.Palpate the chest wall surface to locate the right eighth, ninth, or 10th intercostal spaces between mid and anterior axillary lines. Then place the probe within the zone of apposition, making sure it's entirely situated between the ribs. If a rib appears in the image, adjust the angle of the probe by tilting it up and down until only the diaphragm is visible.On the ultrasound monitor, identify two bright white parallel lines above the liver indicating the pleural and peritoneal membranes. Adjust the depth of the image by clicking on the increase or decrease depth button to maximize the size of the diaphragm. Ensure the diaphragm is visualized in the middle of the monitor.This ensures maximum resolution of the pleural and peritoneal lines from surrounding structures. Next, to optimize image quality, open the ultrasound unit software. Then click the gain button to adjust the image brightness.Increase the gain by clicking the increase button to brighten the image. Conversely, click the decrease button to darken the image. Upon optimizing image quality, click the M mode button to place the ultrasound in the M mode.Position the single vertical scan line between the sections where the pleural and peritoneal lines are the clearest. Run M mode over a full cycle of inspiration and expiration during tidal breathing. Then click on the freeze to capture the actual state and save to save the image.Next, with a skin safe marker, mark the probe's location on the patient's body. Once the examination is complete, click the End Exam button. Next, for image analysis, start the DICOM viewer software and open the necessary DICOM files.Then click on the distance tool and draw a straight line from the inner edge of the pleural membrane to the inner edge of the peritoneal membrane at end expiration. Record this value as diaphragm thickness. Repeat similar measurements at peak inspiration of the same breath to obtain the diaphragm thickness at peak inspiration.Analyze diaphragm thickness at end expiration and peak inspiration from the same breath to assess the diaphragm thickening fraction. Then using the given equation, calculate the diaphragm thickening fraction for each breath. In healthy adults, resting end expiratory diaphragm thickness ranges from 1.5 to 5.0 millimeters, while tidal diaphragm thickening fraction falls between 15 to 30%