This study was supported by the Seoul National University Bundang Hospital Research Fund (14-2015-022) and by the Ministry of Trade Industry &#38; Energy(MOTIE, Korea), Ministry of Science &#38; ICT(MSIT, Korea), and Ministry of Health &#38; Welfare(MOHW, Korea) under Technology Development Program for AI-Bio-Robot-Medicine Convergence (20001650). We would like to thank Su-Bin Park, Nu-Ri Kim and Ye-Lin Jang for helping to prepare and proceed with the video shooting.

All procedures were reviewed and approved by the Seoul National University Bundang Hospital Institutional Review Board (IRB). To recruit healthy participants, posters were used to advertise around the hospital.
1. Experimental set-up
<ol>
	<li>Participant recruitment
	<ol>
		<li>Perform the subject screening process using the following inclusion criteria: 1) healthy, between 18 and 50 years old; 2) right-handed, assessed by Edinburgh handedness inventory14; 3) able to wear the head mount display for VR and to detect objects within VR; and 4) no history of diseases affecting the brain, such as stroke, Parkinson&#39;s disease, or traumatic brain injury. 
		NOTE: These criteria were designed to screen participants with the ability to participate in the experiment and regulate factors affecting the results.</li>
		<li>Recruit participants and provide a detailed explanation of the entire study and expected clinical issues. Consent must be obtained prior to inclusion.</li>
	</ol>
	</li>
	<li>Experimental system 
	NOTE: A customized VPAT system using an immersive VR system and depth-sensing camera was used. Functional infrared spectroscopy (fNIRS) was simultaneously used to investigate the cortical activation. VPAT and fNIRS were linked together for the experiment (Figure 1).
	<ol>
		<li>VPAT system 
		NOTE: The VPAT system consists of a head mount display for VR implementation, a hand tracking sensor that can recognize hand gestures for intuitive input by the user, and a hardware push button. The overall composition is shown in Figure 1.
		<ol>
			<li>Make sure that the hand tracking sensor is not tilted in front of the head mount display.</li>
			<li>Check that the reference camera for the VR system is properly installed on top of the front monitor.</li>
			<li>Secure the push button in a location near the hand to be used by the participant for the experiment.</li>
			<li>Run the software to make sure there are no errors. 
			NOTE: The virtual environment was implemented to match the actual environment as close as possible. The task was performed through hand pointing within the virtual environment and button input through the push button.</li>
		</ol>
		</li>
		<li>fNIRS
		<ol>
			<li>Use a commercial fNIRS system including a personal computer (PC), 31 optodes (15 light sources and 16 detectors), textile EEG caps, and data recording software.</li>
		</ol>
		</li>
		<li>Linkage between VPAT system and fNIRS (Figure 1).
		<ol>
			<li>Use the remote keyboard control software using TCP/IP communication to synchronize the starting event in the VPAT system with the timing of recording in the fNIRS system.</li>
			<li>Use the remote command key in the computer to start fNIRS recording.</li>
		</ol>
		</li>
	</ol>
	</li>
</ol>
2. Experimental set-up (Figure 2)
<ol>
	<li>fNIRS measurement setting
	<ol>
		<li>Place the participant in a chair with his/her back in a straight posture, about fifteen centimeters away from the table. Confirm that the participant&#39;s hand does not hit the table when reaching out.</li>
		<li>For fNIRS cap setting, select the cap size according to the participant&#39;s head circumference. Place the cap so that the vertex (Cz) is located at the intersection of the midpoint between the inion and nasion and the midpoint between the left preauricular and right preauricular areas. Display the montage on the screen and connect 15 sources and 24 detectors to the montage. If necessary to improve the gain from the light source, use conductive gel after hair preparation and insert the optode. Have the participant wear a retaining cap. 
		NOTE: The study used three different sizes of textile EEG caps with circumferences of 54, 56, and 58 cm.</li>
		<li>For software setting (calibration, etc.), run the fNIRS system software and load the neglect montage.</li>
		<li>Let the montage be displayed on the screen and set 15 sources and 24 detectors according to the montage (Figure 3).</li>
		<li>Press the calibrate button. If &#34;Lost&#34; is displayed on the screen, repeat the hair preparation, and then recalibrate.</li>
	</ol>
	</li>
	<li>VPAT system setting
	<ol>
		<li>Connect the HMD, reference camera, and Leap motion camera, and push the button connecting the computer to set up the VPAT system.</li>
		<li>Mount the virtual reality head-mounted display (VR HMD) on the participant&#39;s head over the cap for fNIRS. Make sure to avoid movement of the cap.</li>
		<li>Run the VPAT software. Enter the participant&#39;s information (name abbreviation, age, handedness) and press the &#34;Start&#34; button.</li>
		<li>Confirm the visualization of the virtual hand in the display. Proceed with a two-step calibration (i.e., screen calibration and target distance calibration).</li>
		<li>Instruct the participant to watch the red cross mark (+) in the center, then press the &#34;r&#34; key to calibrate the screen. 
		NOTE: Screen calibration places the virtual space in front of the user&#39;s visual range by recentering the coordinate system.</li>
		<li>Instruct the participant to point to the target (i.e., ball) with his or her right hand, then press the &#34;O&#34; key to calibrate the hand position. 
		NOTE: In our study, the object that the participant had to target was a white ball on a pink stick that came down from the top of the view. Target distance calibration places the target within the reach of the user. This is used to correctly position the target during the experiment.</li>
		<li>After calibration, press the &#34;w&#34; key to begin the experiment.</li>
	</ol>
	</li>
	<li>VPAT and fNIRS linkage setting
	<ol>
		<li>Use the event synchronization software to enter the trigger for analysis into fNIRS and connect VPAT to fNIRS.</li>
		<li>For time synchronization between VPAT and fNIRS, connect the computers with the two systems to the same network, and then synchronize them through the self-produced key transferring program.</li>
		<li>After connecting through the IP and Port inputs of both computers, start the experiment session via the &#34;w&#34; key in the VPAT program. The event synchronization software is executed automatically, and triggers during execution are automatically transferred to fNIRS and saved.</li>
		<li>After the experiment, obtain the software auto-termination and VPAT data. Then stop the VPAT and fNIRS system software. 
		NOTE: The participants must return their hands to their original position after pointing during the VPAT experiment.</li>
	</ol>
	</li>
</ol>
3. Experiment to validate VPAT system
<ol>
	<li>Block designed experiment with fNIRS recording (Figure 4)
	<ol>
		<li>After completing the set-up process in step 2, confirm the participant&#39;s readiness to start the experiment.</li>
		<li>Start the VPAT system without the prism mode and instruct the participant to point to the target in the VR system immediately for familiarization with the procedure.</li>
		<li>Each phase consists of blocks for pointing, clicking, or resting (Figure 4). Again, instruct the participant to click on the button or point to the target in the VR system with their right index finger as quickly as possible.</li>
		<li>Start the experiment with four phases simultaneously with fNIRS recording by clicking the start key. 
		NOTE: During the pointing task, the white ball had to be touched within a fixed time.
		<ol>
			<li>Instruct the participants to point, click, or rest when the appropriate icon appears. 
			NOTE: During the task, pointing and clicking were indicated by an icon directly above the white ball and right side of the timer bar. The time to perform the task was indicated by the timer bar as shown in Figure 2.</li>
			<li>Tell the participant to touch the target that appears on the left or right side within 3 s. For the clicking block, instruct the participant to press the push button. 
			NOTE: The target set containing the white ball was located at a distance of -10&#176; or 10&#176; from the participant&#39;s center, obtained by calibration. The target set appeared randomly on the right or left side. According to the experimental design, the target appeared for 3 s, then disappeared, and then regenerated to a new position.</li>
			<li>Ensure that the participant performs the same way when the phase is switched. 
			NOTE: In the pointing task, Virtual Prism Adaptation Mode showed a deviation of 10&#176; or 20&#176; to the left side of the imaginary hand in the VR space relative to the participant&#39;s head. Zero degrees indicated that the positions of the virtual hand and the actual hand coincided. 
			NOTE: The experiment (Figure 4) consists of a total of four phases, with each phase consisting of pointing and clicking or rest alternately (Phase 1 and 4 were pointing and clicking, and phase 2 and 3 were pointing and resting).</li>
		</ol>
		</li>
	</ol>
	</li>
</ol>
4. Data analysis
<ol>
	<li>Pointing error analysis 
	NOTE: The data were stored from the moment the experimenter pressed the start button &#34;w&#34;. The data were automatically stored at about 60 Hz every frame through the VPAT software. The phase name, elapsed time, and virtual index finger position were stored over time. The pointing error was the angle value between the target and index finger, centered on the participant&#39;s head position.
	<ol>
		<li>Classify the pointing task data by phases (pre-VPAT, VPAT 10&#176;, VPAT 20&#176;, post-VPAT).</li>
		<li>Classify the data of the pointing task and the clicking task in the data of each phase (phase 1 and 4).</li>
		<li>Classify the data by sub-phase in units of 30 s according to each phase and each type of task.</li>
		<li>Extract the median value of 10 trial error (pointing error) values from the index finger position data for median pointing error analysis.</li>
		<li>Use the repeated measures analysis of variance test (ANOVA) to analyze the difference between each phase. 
		NOTE: In the case of hand tracking using the Leap motion sensor, outliers were due to occlusion or false detection of the hand posture. With the exception of false hand position data, the median value was used to find the representative pointing error value in the sub-phase.</li>
	</ol>
	</li>
	<li>fNIRS data processing
	<ol>
		<li>Launch the fNIRS analysis software and load the raw data file and probe information.</li>
		<li>Perform a marker setting process by editing the event record to verify each condition during the experiment.</li>
		<li>Carry out data preprocessing by deleting the experimentally irrelevant time intervals, remove artifacts, such as steps and spikes, and apply frequency filters to exclude experimentally irrelevant frequency bands. 
		NOTE: All data sets were filtered with a 0.01 Hz high-pass filter and a 0.2 Hz low-pass filter to remove instrumental or physiological noise contributions.</li>
		<li>Specify wavelengths by entering the value of the peak illumination wavelengths (i.e., 760 and 850 nm). Use a physical distance of 3 cm between the source and detector for channel.</li>
		<li>Select the baseline field, which refers to the time period that corresponds to a baseline wherein participants are typically resting quietly. 
		NOTE: We selected the baseline field as the full-time course of the data set, which was the default setting.</li>
		<li>Compute the time series of hemodynamic states to finish the preprocessing from the filtered data.</li>
	</ol>
	</li>
</ol>

<ol>
	<li>Appelros, P., Karlsson, G. M., Seiger, A., Nydevik, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neglect+and+anosognosia+after+first-ever+stroke:+incidence+and+relationship+to+disability.">Neglect and anosognosia after first-ever stroke: incidence and relationship to disability.</a> Journal of Rehabilitation Medicine. 34 (5), 215-220 (2002).</li><li>Buxbaum, 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=Hemispatial+neglect+subtypes,+neuroanatomy,+and+disability.">Hemispatial neglect subtypes, neuroanatomy, and disability.</a> Neurology. 62 (5), 749-756 (2004).</li><li>Jehkonen, 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=Visual+neglect+as+a+predictor+of+functional+outcome+one+year+after+stroke.">Visual neglect as a predictor of functional outcome one year after stroke.</a> Acta Neurologica Scandinavica. 101 (3), 195-201 (2000).</li><li>Jehkonen, M., Laihosalo, M., Kettunen, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Impact+of+neglect+on+functional+outcome+after+stroke&#8211;a+review+of+methodological+issues+and+recent+research+findings.">Impact of neglect on functional outcome after stroke&#8211;a review of methodological issues and recent research findings.</a> Restorative Neurology and Neuroscience. 24 (4-6), 209-215 (2006).</li><li>Mizuno, 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=Prism+adaptation+therapy+enhances+rehabilitation+of+stroke+patients+with+unilateral+spatial+neglect:+a+randomized,+controlled+trial.">Prism adaptation therapy enhances rehabilitation of stroke patients with unilateral spatial neglect: a randomized, controlled trial.</a> Neurorehabilitation and Neural Repair. 25 (8), 711-720 (2011).</li><li>Shiraishi, H., Yamakawa, Y., Itou, A., Muraki, T., Asada, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Long-term+effects+of+prism+adaptation+on+chronic+neglect+after+stroke.">Long-term effects of prism adaptation on chronic neglect after stroke.</a> NeuroRehabilitation. 23 (2), 137-151 (2008).</li><li>Yang, N. Y., Zhou, D., Chung, R. C., Li-Tsang, C. W., Fong, K. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Rehabilitation interventions for unilateral neglect after stroke: a systematic review from 1997 through 2012. , (2013).</li><li>Rossetti, 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=Prism+adaptation+to+a+rightward+optical+deviation+rehabilitates+left+hemispatial+neglect.">Prism adaptation to a rightward optical deviation rehabilitates left hemispatial neglect.</a> Nature. 395 (6698), 166-169 (1998).</li><li>Barrett, A., Goedert, K. M., Basso, 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=Prism+adaptation+for+spatial+neglect+after+stroke:+translational+practice+gaps.">Prism adaptation for spatial neglect after stroke: translational practice gaps.</a> Nature Reviews Neurology. 8 (10), 567-577 (2012).</li><li>Maxton, C., Dineen, R., Padamsey, R., Munshi, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Don't+neglect+'neglect'-an+update+on+post+stroke+neglect.">Don't neglect 'neglect'-an update on post stroke neglect.</a> International Journal of Clinical Practice. 67 (4), 369-378 (2013).</li><li>Gammeri, R., Turri, F., Ricci, R., Ptak, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Adaptation+to+virtual+prisms+and+its+relevance+for+neglect+rehabilitation:+a+single-blind+dose-response+study+with+healthy+participants.">Adaptation to virtual prisms and its relevance for neglect rehabilitation: a single-blind dose-response study with healthy participants.</a> Neuropsychol Rehabilitation. , 1-14 (2018).</li><li>Saj, A., Cojan, Y., Assal, F., Vuilleumier, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Prism+adaptation+effect+on+neural+activity+and+spatial+neglect+depend+on+brain+lesion+site.">Prism adaptation effect on neural activity and spatial neglect depend on brain lesion site.</a> Cortex. 119, 301-311 (2019).</li><li>Redding, G. M., Wallace, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Generalization+of+prism+adaptation.">Generalization of prism adaptation.</a> Journal of Experimental Psychology: Human Perception and Performance. 32 (4), 1006-1022 (2006).</li><li>Caplan, B., Mendoza, J. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Encyclopedia of Clinical Neuropsychology. , 928 (2011).</li><li>Kim, W. S., Paik, N. J., Cho, 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> 2017 International Conference on Virtual Rehabilitation (ICVR). , 1-2 (2017).</li><li>Munafo, J., Diedrick, M., Stoffregen, T. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+virtual+reality+head-mounted+display+Oculus+Rift+induces+motion+sickness+and+is+sexist+in+its+effects.">The virtual reality head-mounted display Oculus Rift induces motion sickness and is sexist in its effects.</a> Experimental Brain Research. 235 (3), 889-901 (2017).</li><li>Merhi, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Proceedings of the Human Factors and Ergonomics Society Annual Meeting. , 2618-2622 (2018).</li><li>Duh, H. B. L., Parker, D. E., Furness, T. 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> Proceedings of 9th International Conference on Human-Computer Interaction. , 5-10 (2018).</li><li>Prothero, J. D., Draper, M. 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Won-Seok Kim, Sungmin Cho, and Nam-Jong Paik have a patent entitled &#34;Method, system and readable recording medium of creating visual stimulation using virtual model&#34;, number 10-1907181, which is relevant to this work.

This study implemented the prism adaption therapy using a translated hand movement in a VR environment. It investigated whether the deviation implemented was causing angle overshooting and behavioral adaptation, as in conventional prism adaption therapy.
In the median pointing error result (Figure 5) and the first pointing error result, the pointing error changed significantly when the phase was switched. Although some hand-recognition errors were eliminated, there may still be false detection. The use of a median value to eliminate systematic error, such as false tracking, showed that the average pointing error results were lower than expected. Post-prismatic adaptation was constantly shown in each subject (Figure 5B). These results showed similar behavioral adaptation to the conventional prism adaptation therapy.
There were some problems in the experiment. False detection of the hand occurred frequently in the pointing task. In some cases, even though the hand reached the target during pointing, the virtual hand was not tracked due to a Leap motion recognition error. In addition, because the participants were wearing HMD in the clicking task, it was difficult for them to locate the push button and the experimenter had to provide continuous assistance. The weight of the HMD and its long-term application could also cause pain in the area that comes into contact with the fNIRS optode. Therefore, there were times when the HMD was lifted or the participants themselves were holding the HMD.
If we overcome the shortcomings of the system and consolidate the results of the experiment through more data analysis, including fNIRS data, it could potentially be used in the treatment of visuospatial neglect. In addition, game-friendly contents can be applied to present an immersive and fun treatment modality. Nonetheless, further study with a more advanced VPAT system proving clinical efficacy in stroke patients with visuospatial neglect is needed.
Several previous studies have reported motion sickness induced by the use of Immersive VR, or head-mounted VR sets16. Motion sickness is reported to be infrequent if VR is implemented in seated positions17. Motion mismatch can also cause motion sickness, but it can be reduced by independently configuring the background in the virtual environment18,19. In this system, only the hand deviation angle caused motion mismatch, which should have less impact on motion sickness overall.
Participants in this experiment were normal adults, so there were no consistent problems. However, to be used as therapeutic treatment for stroke patients, the above issues need to be considered, and virtual prism therapy protocols need to be taken into account, such as taking breaks during treatment or the length of treatment time.

Hemispatial neglect, which affects the ability to perceive the contralateral hemispatial visual field, is a common impairment after stroke1,2. Although rehabilitation after hemispatial neglect is important, due to its association with poor functional and social outcomes, rehabilitation is often underutilized in real clinical practice3,4.
Among the various existing rehabilitation approaches suggested for hemispatial neglect, prism adaptation (PA) therapy has proven effective for recovery and improvement in hemispatial neglect in patients with subacute or chronic stroke5,6,7,8. However, conventional PA is underutilized due to several drawbacks9,10. These include 1) high cost and time requirement due to the prism lens needing to be changed to adjust to the degree of deviation; 2) the need to set up additional materials to be pointed at and to mask the hand trajectory; and 3) PA can only be used by patients who can sit and control their head position.
A recent study reproducing the adaptation effects in the virtual reality (VR) environment reported that it may be possible for the virtual prism adaptation therapy (VPAT) to have different effects depending on the subtypes of neglect11. It was also suggested that cortical activation for PA may vary according to brain lesions12. However, little is known about the cortical activation pattern seen in VR-induced PA.
To overcome these obstacles and promote the use of PA in a clinical setting, we developed a new PA therapy system using an immersive VR technology called virtual prism adaptation therapy (VPAT), via the use of a depth-sensing camera. We designed an immersive VR system with the ability to provide visual feedback about the position of a virtual limb to promote spatial realignment13. Using this immersive VR technology, which mimicked the effect of conventional PA, we designed an experiment to validate the VPAT system in healthy participants.
By conducting our visualized experimental protocol, we investigated whether the new VPAT system can induce behavioral adaptation, similar to conventional PA. Additionally, we would like to explore whether the VPAT system can induce the activation in the cortical regions associated with visuospatial perception or recovery of hemispatial neglect after stroke.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>EASYCAP</td>
			<td>Easycap</td>
			<td>C-SAMS</td>
			<td>Platform to accommodate fNIRS optodes</td>
		</tr>
		<tr>
			<td>Leap Motion 3D Motion Controller</td>
			<td>Ultrahaptics</td>
			<td>FBA_LM-C01-US</td>
			<td>Hand detection device attached HMD</td>
		</tr>
		<tr>
			<td>Leap Motion VR Developer Mount for VR Headset</td>
			<td>Ultrahaptics</td>
			<td>VR-UAZ</td>
			<td></td>
		</tr>
		<tr>
			<td>Matlab R2015a</td>
			<td>Mathworks</td>
			<td></td>
			<td>Programming language running with NIRStar</td>
		</tr>
		<tr>
			<td>NIRScout</td>
			<td>Medical Technology LLC</td>
			<td>NSC-CORE</td>
			<td>fNIRS system</td>
		</tr>
		<tr>
			<td>nirsLAB v201605</td>
			<td>Medical Technology LLC</td>
			<td></td>
			<td>Software for analyzing data collected with NIRScout</td>
		</tr>
		<tr>
			<td>NIRStar 14.1</td>
			<td>Medical Technology LLC</td>
			<td></td>
			<td>NIRScout Acquisition Software</td>
		</tr>
		<tr>
			<td>Occulus Rift DK2</td>
			<td>Occulus</td>
			<td></td>
			<td>VR HMD</td>
		</tr>
		<tr>
			<td>PowerMate USB Multimedia Controller</td>
			<td>Griffin Technology</td>
			<td>NA16029</td>
			<td>Push Button in task</td>
		</tr>
		<tr>
			<td>SuperLab 5.0</td>
			<td>Cedrus Corp.</td>
			<td></td>
			<td>Synchronize the stimulus presentations allied to NIRScout</td>
		</tr>
	</tbody>
</table>

virtual prism adaptation therapy: protocol for validation in healthy adults

Hemispatial neglect is a common impairment after stroke. It is associated with poor functional and social outcomes. Therefore, an adequate intervention is imperative for the successful management of hemispatial neglect. However, the clinical use of various interventions is limited in real clinical practice. Prism adaptation therapy is one of the most evidence-based rehabilitation modalities to treat hemispatial neglect. To overcome any possible shortcoming that may occur with prism therapy, we developed a new system using immersive virtual reality and depth-sensing camera to create a virtual prism adaptation therapy (VPAT). To validate the VPAT system, we designed an experimental protocol investigating the behavioral errors and changes in cortical activation via the VPAT system. Cortical activation was measured by functional near infrared spectroscopy (fNIRS). The experiment consisted of four phases. All four included clicking, pointing or rest applied to right-handed healthy people. Clicking versus pointing was used for investigating the cortical region related with the gross motor task, and pointing with VPAT versus pointing without VPAT was used for investigating the cortical region associated with visuospatial perception. The preliminary results from four healthy participants showed that pointing errors by the VPAT system was similar to the conventional prism adaptation therapy. Further analysis with more participants and fNIRS data, as well as a study in patients with stroke may be required.

Data from four healthy participants (1 man and 3 women) were used as representative results. A pointing error is shown in Figure 5A, with the averages of median value of 10 trials in the sub-phase of each pointing task lasting 30 s. Values on average for the median pointing errors in the first block of each phase were 0.45 &#177; 0.92 (pre-VPAT), 4.69 &#177; 3.08 (VPAT 10&#176;), 5.43 &#177; 2.22 (VPAT 20&#176;), and -5.17 &#177; 1.60 (post-VPAT). The trend of pointing error change was statistically significant (p = 0.001) via the repeated measures ANOVA. A pointing error for each subject is presented in Figure 5B, illustrating the adaptation during the VPAT phase and post-prismatic adaptation (negative pointing error).
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/60639/60639fig01.jpg" /> 
Figure 1: Experimental setting with VPAT and fNIRS linkage system. VPAT = virtual prism adaptation therapy; fNIRS = functional near infrared spectroscopy. This figure was previously published by Kim et al.15 <a href="https://www.jove.com/files/ftp_upload/60639/60639fig01large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/60639/60639fig02.jpg" /> 
Figure 2: The subject performing the experiment with VPAT and fNIRS system. VPAT = virtual prism adaptation therapy; fNIRS = functional near infrared spectroscopy. <a href="https://www.jove.com/files/ftp_upload/60639/60639fig02large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/60639/60639fig03.jpg" /> 
Figure 3: Montage containing 54 channels by arranging 15 light sources (red circles) and 24 detectors (blue circles) at intervals of 3 cm. Space between the nearest sources and the detector constituted one channel, which is represented as yellow circles with a number. <a href="https://www.jove.com/files/ftp_upload/60639/60639fig03large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/60639/60639fig04.jpg" /> 
Figure 4: Experimental design. VPAT = virtual prism adaptation therapy; Pt = pointing; Cl = clicking; Re = resting. <a href="https://www.jove.com/files/ftp_upload/60639/60639fig04large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 5" class="xfigimg" src="/files/ftp_upload/60639/60639fig05.jpg" /> 
Figure 5: Pointing errors in each block. (A) Average value graph of subject&#39;s median pointing error in each block. This figure was previously published by Kim et al.15 (B) Median pointing error in each block by each subject. The counterclockwise direction (i.e. left from the target) is the positive value. <a href="https://www.jove.com/files/ftp_upload/60639/60639fig05large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>

Watch this Scientific Journal Video about Virtual Prism Adaptation Therapy: Protocol for Validation in Healthy Adults at JoVE.com

Virtual Prism Adaptation Therapy: Protocol for Validation in Healthy Adults

This experimental protocol demonstrates the use of virtual prism adaptation therapy (VPAT) in healthy adults and the association between VPAT and functional near infrared spectroscopy to determine the effect of VPAT on cortical activation. Results suggest that VPAT may be feasible and could induce similar behavioral adaptation as conventional prism adaptation therapy.

Hemispatial neglect is a common impairment after stroke. It is associated with poor functional and social outcomes. Therefore, an adequate intervention is ...

virtual-prism-adaptation-therapy-protocol-for-validation-healthy

Nanyang Technological University

Research

JoVE Journal

Medicine

6.7K Views.  Delvine Inc.. This experimental protocol demonstrates the use of virtual prism adaptation therapy (VPAT) in healthy adults and the association between VPAT and functional near infrared spectroscopy to determine the effect of VPAT on cortical activation. Results suggest that VPAT may be feasible and could induce similar behavioral adaptation as conventional prism adaptation therapy.

Video: Virtual Prism Adaptation Therapy: Protocol for Validation in Healthy Adults

Production and Detection of Reactive Oxygen Species (ROS) in Cancers

Reactive oxygen species include a number of molecules that damage DNA and RNA and oxidize proteins and lipids (lipid peroxydation). These reactive molecules contain an oxygen and include H2O2 (hydrogen peroxide), NO (nitric oxide), O2- (oxide anion), peroxynitrite (ONOO-), hydrochlorous acid (HOCl), and hydroxyl radical (OH-).
Oxidative species are produced not only under pathological situations (cancers, ischemic/reperfusion, neurologic and cardiovascular pathologies, infectious diseases, inflammatory diseases 1, autoimmune diseases 2, etc&hellip;) but also during physiological (non-pathological) situations such as cellular metabolism 3, 4. Indeed, ROS play important roles in many cellular signaling pathways (proliferation, cell activation 5, 6, migration 7 etc..). ROS can be detrimental (it is then referred to as "oxidative and nitrosative stress") when produced in high amounts in the intracellular compartments and cells generally respond to ROS by upregulating antioxidants such as superoxide dismutase (SOD) and catalase (CAT), glutathione peroxidase (GPx) and glutathione (GSH) that protects them by converting dangerous free radicals to harmless molecules (i.e. water). Vitamins C and E have also been described as ROS scavengers (antioxidants).
Free radicals are beneficial in low amounts 3. Macrophage and neutrophils-mediated immune responses involve the production and release of NO, which inhibits viruses, pathogens and tumor proliferation 8. NO also reacts with other ROS and thus, also has a role as a detoxifier (ROS scavenger). Finally NO acts on vessels to regulate blood flow which is important for the adaptation of muscle to prolonged exercise 9, 10. Several publications have also demonstrated that ROS are involved in insulin sensitivity 11, 12.
Numerous methods to evaluate ROS production are available. In this article we propose several simple, fast, and affordable assays; these assays have been validated by many publications and are routinely used to detect ROS or its effects in mammalian cells. While some of these assays detect multiple ROS, others detect only a single ROS.

Production and Detection of Reactive Oxygen Species ROS in Cancers

Here we propose simple methods to test and evaluate the presence of reactive oxygen species in cells.

Reactive oxygen species include a number of molecules that damage DNA and RNA and oxidize proteins and lipids (lipid peroxydation). These reactive ...

Development of an Audio-based Virtual Gaming Environment to Assist with Navigation Skills in the Blind

Audio-based Environment Simulator (AbES) is virtual environment software designed to improve real world navigation skills in the blind. Using only audio based cues and set within the context of a video game metaphor, users gather relevant spatial information regarding a building's layout. This allows the user to develop an accurate spatial cognitive map of a large-scale three-dimensional space that can be manipulated for the purposes of a real indoor navigation task. After game play, participants are then assessed on their ability to navigate within the target physical building represented in the game. Preliminary results suggest that early blind users were able to acquire relevant information regarding the spatial layout of a previously unfamiliar building as indexed by their performance on a series of navigation tasks. These tasks included path finding through the virtual and physical building, as well as a series of drop off tasks. We find that the immersive and highly interactive nature of the AbES software appears to greatly engage the blind user to actively explore the virtual environment. Applications of this approach may extend to larger populations of visually impaired individuals.

Audio-based Environment Simulator (AbES) is virtual environment software designed to improve real world navigation skills in the blind.

Audio-based Environment Simulator (AbES) is virtual environment software designed to improve real world navigation skills in the blind. Using only audio ...

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 ...

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 ...

A Method for Quantifying Upper Limb Performance in Daily Life Using Accelerometers

A key reason for referral to rehabilitation services after stroke and other neurological conditions is to improve one's ability to function in daily life. It has become important to measure a person's activities in daily life, and not just measure their capacity for activity in the structured environment of a clinic or laboratory. A wearable sensor that is now enabling measurement of daily movement is the accelerometer. Accelerometers are commercially-available devices resembling large wrist watches that can be worn throughout the day. Data from accelerometers can quantify how the limbs are engaged to perform activities in peoples' homes and communities. This report describes a methodology to collect accelerometry data and turn it into clinically-relevant information. First, data are collected by having the participant wear two accelerometers (one on each wrist) for 24 h or longer. The accelerometry data are then downloaded and processed to produce four different variables that describe key aspects of upper limb activity in daily life: hours of use, use ratio, magnitude ratio, and the bilateral magnitude. Density plots can be constructed that visually represent the data from the 24 h wearing period. The variables and their resultant density plots are highly consistent in neurologically-intact, community-dwelling adults. This striking consistency makes them a useful tool for determining if upper limb daily performance is different from normal. This methodology is appropriate for research studies investigating upper limb dysfunction and interventions designed to improve upper limb performance in daily life in people with stroke and other patient populations. Because of its relative simplicity, it may not be long before it is also incorporated in clinical neurorehabilitation practice.

This protocol describes a method to quantify upper limb performance in daily life using wrist-worn accelerometers.

A key reason for referral to rehabilitation services after stroke and other neurological conditions is to improve one's ability to function in daily life. ...

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 ...

Home-Based Transcranial Direct Current Stimulation Device Development: An Updated Protocol Used at Home in Healthy Subjects and Fibromyalgia Patients

Transcranial direct current stimulation (tDCS) is a non-invasive brain stimulation (NIBS) method, which modulates the membrane potential of neurons in the cerebral cortex by a low-intensity direct current. tDCS is a low-cost technique with minimal adverse effects and easy application. This neurostimulation method has a promising future to improve pain therapy, treatment of neuropsychiatric disorders, and physical rehabilitation.
Current studies demonstrate the benefits of using tDCS over consecutive multiple sessions. However, the daily displacement to the specialized centers, travel costs, and disruptions to daily activities are some of the difficulties faced by patients. Thus, to be more comfortable, easy-to-use, and not disrupt daily commitments, a home-based tDCS was designed. Therefore, the objective of this study was to evaluate the feasibility of a portable tDCS device for home use in healthy subjects and fibromyalgia patients.
Despite increased tDCS use and a reasonably large body of research on the effects across a range of clinical conditions, there is a limited amount of research on developing secure devices that guarantee the dose and contain a block system to avoid excessive use. Therefore, we used a tDCS device with a security system to permit daily use for 20 minutes with a minimal interval of 12 hours between sessions. A programmer preconfigures the equipment, which has a neoprene cap that allows the electrode positions in any assembly, according to individualized protocols for treatments or research. After, researchers can assess the effectiveness of treatment, and its adherence using information kept in the device software.
Results suggest that the device is feasible for home use, with proper monitoring of adherence and contact impedance. There were reports of a few adverse effects, which do not differ from those reported in the literature in studies with the treatment under direct supervision.

This study provides an updated home-based tDCS protocol that enables subjects to receive the beneficial effects of tDCS at home with an easy to use device with settings to control the use and dosage, enhancing the feasibility for long-term use at home.

Transcranial direct current stimulation (tDCS) is a non-invasive brain stimulation (NIBS) method, which modulates the membrane potential of neurons in the ...

Fat-Water Phantoms for Magnetic Resonance Imaging Validation: A Flexible and Scalable Protocol

As new techniques are developed to image adipose tissue, methods to validate such protocols are becoming increasingly important. Phantoms, experimental replicas of a tissue or organ of interest, provide a low cost, flexible solution. However, without access to expensive and specialized equipment, constructing stable phantoms with high fat fractions (e.g., &#62;50% fat fraction levels such as those seen in brown adipose tissue) can be difficult due to the hydrophobic nature of lipids. This work presents a detailed, low cost protocol for creating 5x 100 mL&#160;phantoms with fat fractions of 0%, 25%, 50%, 75%, and 100% using basic lab supplies (hotplate, beakers, etc.) and easily accessible components (distilled water, agar, water-soluble surfactant, sodium benzoate, gadolinium-diethylenetriaminepentacetate (DTPA) contrast agent, peanut oil, and oil-soluble surfactant). The protocol was designed to be flexible; it can be used to create phantoms with different fat fractions and a wide range of volumes. Phantoms created with this technique were evaluated in the feasibility study that compared the fat fraction values from fat-water magnetic resonance imaging to the target values in the constructed phantoms. This study yielded a concordance correlation coefficient of 0.998 (95% confidence interval: 0.972-1.00). In summary, these studies demonstrate the utility of&#160;fat phantoms for validating adipose tissue imaging techniques across a range of clinically relevant tissues and organs.

The purpose of this work is to describe a protocol for creating a practical fat-water phantom that can be customized to produce phantoms with varying fat percentages and volumes.

As new techniques are developed to image adipose tissue, methods to validate such protocols are becoming increasingly important. Phantoms, experimental ...

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 ...

Clinical Practice Protocol of Creative Music Therapy for Preterm Infants and Their Parents in the Neonatal Intensive Care Unit

Creative music therapy for preterm infants and their parents (CMT) has emerged as a promising family-integrated early intervention involving communicative musicality to improve infant development, parental well-being, and bonding. It aims at relaxing and nurturing the infant as well as promoting safety and social interaction for the parent-infant dyad. A music therapist specially trained in CMT hums or sings in an infant-directed, improvised, lullaby style continually adjusting to the individual needs, expressions, and breathing pattern of the preterm infant. Based on the principles of family-integrated care, the family is incorporated individually in the therapeutic process, namely by delivering CMT during kangaroo care (KC) and by motivating and facilitating parental vocal interaction with their infant to strengthen the parent-infant bonding. CMT aims at relaxing, stimulating, and coregulating premature infants at a time when many other interventions are still risky and can overwhelm the vulnerable patient group. CMT may be advantageous not by educating and teaching parents, but rather by uncovering the intuitive capacities of parenting that are often overshadowed by the traumatic experience of preterm birth. However, CMT can only be provided when the infants are clinically stable. CMT with parental integration is feasible when parents are available and receptive to participate. This paper presents a detailed protocol on how to use CMT to empower preterm infants and their families.

Creative music therapy for preterm infants and their parents has emerged as a promising family-integrated early intervention. We present a detailed protocol on how to use vocal interaction, humming, or singing to empower preterm infants and their families.

Creative music therapy for preterm infants and their parents (CMT) has emerged as a promising family-integrated early intervention involving communicative ...