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Abstract
Introduction Upper limb (UL) impairment is common in people with multiple sclerosis (pwMS), and functional recovery of the UL is a key rehabilitation goal. Technology-based approaches, like virtual reality (VR), are increasingly promising. While most VR environments are task-oriented, our clinical approach integrates neuroproprioceptive ‘facilitation and inhibition’ (NFI) principles. To advance this, we developed immersive VR software based on NFI principles targeting UL function and sit-to-stand ability. This study aims to evaluate the effectiveness of this VR therapy compared with conventional NFI-based physical therapy in pwMS. Our study uniquely applies advanced imaging techniques, along with biological molecular assessments, to explore adaptive processes induced by VR rehabilitation.
Methods and analysis This double-arm, randomised, assessor-blinded, controlled trial runs over 2 months (1 hour, 2 times per week). PwMS with mild to severe disability will receive either VR therapy or real-world physical therapy. Primary outcomes include the nine-hole peg test, box and block test, handgrip strength, tremor and five times sit-to-stand test. Secondary measures include the Multiple Sclerosis Impact Scale, the 5-level EQ-5D questionnaire and kinematic analysis. Adaptive processes will be monitored using imaging techniques (functional MRI and tractography), molecular genetic methods (long non-coding RNAs) and immune system markers (leukocytes, dendritic cells). The International Classification of Functioning, Disability and Health brief set for MS will map the bio-psycho-social context of participants.
Ethics and dissemination This project and its amendments were approved by the Ethics Committee of the Institute for Clinical and Experimental Medicine and Thomayer Hospital (1983/21+4772/21 (G-21–02) and the Ethics Committee of Kralovske Vinohrady University Hospital (EK-VP/38/0/2021) in Prague, Czechia (with single enrolment). The findings of this project will be disseminated through scientific publications, conferences, professional networks, public engagement, educational materials and stakeholder briefings to ensure a broad impact across clinical, academic and public domains.
Trial registration number clinicaltrials.gov (NCT04807738).
- Multiple sclerosis
- Virtual Reality
- REHABILITATION MEDICINE
- Neuroradiology
- Functional Magnetic Resonance Imaging
- IMMUNOLOGY
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- Multiple sclerosis
- Virtual Reality
- REHABILITATION MEDICINE
- Neuroradiology
- Functional Magnetic Resonance Imaging
- IMMUNOLOGY
STRENGTHS AND LIMITATIONS OF THIS STUDY
An innovative serious game Virtual Reality (VR) software precisely emulating rehabilitation based on neuroproprioceptive ‘facilitation, inhibition’ principles.
Original approach to physiotherapy fusing VR, and physical therapy methods to facilitate physiological movement of impaired upper extremity function and sit-to-stand transitions.
Methodically near-identical physiotherapy content with VR enables identifying the contribution of the tested software on the final effect on the selected outcome measures.
Outcome measures include advanced functional MRI and tractography evaluating the brain functional connectivity and plasticity triggered by physical therapy intervention.
Unique exploratory assessment with molecular biomarkers of long non-coding RNA, peripheral blood leucocytes and dendritic cell analysis to document the neurobiological impact of the therapy.
Introduction
Upper limb (UL) mobility dysfunction affects 60%–75% of people with multiple sclerosis (PwMS), resulting in reduced participation in activities of daily living.1 2 It has been documented that physical therapy (PT) brings important physical and psychological benefits. PT plays a key role in coping and optimising functional ability3 and improvement of activity level.4
Technologies such as virtual reality (VR) might be of benefit in rehabilitation,5 but only a few studies focused on ULs in PwMS. Those indicated improvement in manual dexterity, coordination, precision, execution time and gross manual dexterity.2 6 7
The current use of VR in PT is mainly based on the principles of sensorimotor learning. Game mechanisms focus on concentration and motivation, aiming at increased task performance repetitions.8 9 Multisensory stimulation entrains the mirror neuron system,10 and stored motor plans are activated, potentiating motor performance11 and producing whole-body illusions.10 VR through multifactorial stimulation influences dopamine centres in the brain.12 Our team has developed a VR rehabilitation programme based on NFI physiotherapeutic methods. Systematic repetition induces a substantial cortical network reorganisation topographically closely related to the trained movement causing synaptogenesis.8 13 NFI-based PT enhances synaptic connections among neurons forming functional networks evocating movement. A suitable combination of afferent stimuli repeatedly activates motor programmes at the subcortical level, inducing adaptive and plastic processes.14 Brain plasticity changes following VR have been documented in poststroke people.15–17 NFI approach affects brain structures involved in motor control,14 18–20 inducing structural changes.19 21 Combining NFI and sensorimotor learning in VR has the potential to further induce adaptive and plastic processes. This is the first study to map the effect of VR PT using functional MRI (fMRI) and magnetic resonance tractography (MRT) in PwMS.
Molecular biomarkers could indicate improvements that support repair or compensatory strategies. Long non-coding RNAs (lncRNAs) have the potential to serve as a biomarker of MS activity degree, its progression phase and the response to disease-modifying drugs.22 High expression of lncRNA MALAT1 inhibits neuronal apoptosis and lowers the proinflammatory interleukin 6 (IL-6).23 High expression of lncRNA MEG3 contributes to the sustained proinflammatory state of microglial cells.24 LncRNA H19 induces microglial apoptosis.25 LncRNA GAS5 and miRNA miR-137 gene expression changes suggest a key regulatory role in MS pathogenesis.26 Our group discovered lncRNA PARTICLE that takes part in the regulation of gene imprinting, epigenetic chromatin modification,27 transcription interference and nuclear export.28 29 We have identified selected lncRNAs (eg, MALAT1, MEG3, H19, GAS5, PARTICLE, TINA, etc) as suitable blood biomarkers for monitoring the efficacy of the intervention.
At the cellular level, this study analyses the composition of peripheral blood leucocytes Natural Killer (NK) cells, monocytes, T cells, B cells and their subpopulations) and the functional condition of generated monocyte-derived dendritic cells (MoDCs) by determining the expression of relevant surface molecules (see methods). The strongest genetic risk factor in MS is HLA-DRB1*15:01 allele encoding the beta chain of HLA (human leukocyte antigen)-DR molecule.30 The research on the determination of dendritic cell (DC) populations is ongoing.31 The conventional DCs play a pivotal role in the pathogenesis of MS32 33 by cooperating with the autoreactive T cells.34 The demyelination process is supported by the microglia and MoDCs.35 The conventional DCs in the relapsing-remitting MS or the secondary progressive MS had a higher proportion of CD40 expression compared with the healthy controls.36 In MS patients, DC frequency and maturity may correlate with disease stage and clinical presentation, but it has not been studied in the context of PT intervention.
Our team proposes a double-arm randomised controlled trial (RCT) that optimises and enhances VR-based PT PwMS with impaired upper extremity function, trunk coordination or transitions from a sitting position. We hypothesise that the use of VR in PT will have a higher effect on clinical functions than identical neuroproprioceptive ’facilitation, inhibition’ (NFI) PT without VR. Moreover, plastic and adaptive processes will be monitored using imaging methods fMRI, MRT and molecular genetic methods (long non-coding RNAs) and monitoring of immune system (the activation of DCs).
Methods and analysis
Study design
This study is designed as a single-blinded RCT with parallel assignment, registered under ID: NCT04807738 in clinical trials.gov. Participants will be randomly assigned into two groups of outpatient rehabilitation programme focusing on ULs, trunk control, stability in sitting and sit-to-stand ability. Both groups will receive physiotherapy based on the NFI principles. The intervention group engages in the VR with active involvement of the physiotherapist, while the control group participates in the same activities without the use of VR. Examination at baseline (E1) and after completion of the PT programme (E2) will consist of clinical examination assessing UL function and sit-to-stand transitions, fMRI, MRT and blood sample analysis.
Study setting
This study will be conducted at the Department of Rheumatology and Physiotherapy, Third Faculty of Medicine, Charles University and Thomayer University Hospital, Czech Republic and Department of Rehabilitation Medicine, Third Faculty of Medicine, Charles University, Prague, Czechia. Both workplaces are equipped with HTC Vive VR devices and tracking devices required for the VR-based PT. The therapeutic VR software for rehabilitation was developed and is further managed by the computer science researchers at the University of West Bohemia, Czech Republic.
Research and management team
The study team includes a coordinator, physiotherapists, occupational therapists (OTs), software developers, molecular biologists, MRI specialists and a statistician. The coordinator manages participant recruitment, eligibility assessment, allocation, assignment and study flow while providing participants with detailed study information. All personnel involved in assessments are blinded to treatment allocation. A blinded OT assessor conducts clinical measurements and administers questionnaire surveys. Molecular biologists handle the processing of peripheral blood samples, including peripheral blood mononuclear cell (PBMC) extraction and lncRNA expression analysis from in-vitro cultivated DCs. Blinded MRI specialists oversee fMRI examinations and data analysis. Four physiotherapists, specifically trained in the NFI-based PT method and certified in this technique, conduct both arms of the intervention. These physiotherapists, as they have an average of 2 years of clinical experience, work under the supervision of an experienced physiotherapist who developed the methodology (KR). The statistician prepares the dataset, will conduct regular audits, ensure data quality, and determine the appropriateness of parametric or non-parametric testing for significance analyses.
Participants
Sample size
Due to the limited number of PwMS patients, the planned number of participants is 70 (35 per group). This sample size is sufficient to detect a difference in the nine hole peg test (NHPT) and in other quantitative outcome measures (OMs) between the intervention and control group with an effect size of Cohen’s d=0.68 (a medium-to-large effect size), a power of 80% and a significance level of 5%, when using the two-tailed independent samples t-test. Expecting 10% of patients not meeting the inclusion criteria and a drop-out rate of 20%, 98 participants should be recruited to achieve the sample size of 70. The power analysis (pwr package37) in R software was used for sample size calculation.
Patient recruitment
Patient recruitment is in cooperation with MS centres in Prague. The study is also advertised in local patient organisations in Prague (Unie Roska). Recruitment of the patients is continuous. All participants will be referred to the coordinator. The recruitment period is from April 2021 to January 2025.
Eligibility criteria
The inclusion criteria are age >18 and <75 years, diagnosis of multiple sclerosis according to the 2017 revised Mc Donald criteria,38 an Expanded Disability Status Scale (EDSS) score ≥2 a ≤7 (27) (determined by a neurologist in the latest patient record) and in the past 3 months prior to recruitment no history of relapse, change in disease-modifying treatment nor corticosteroid therapy.
Exclusion criteria include other factors influencing mobility (history of stroke, pregnancy, traumatic injury of limb/s, severe cardiovascular or orthopaedic dysfunction or impaired cognitive functions during examination and/or consecutive therapy).
The following data will be collected on patient population characteristics: age, sex, weight and height, laterality,39 EDSS, disease duration, MS therapy, use of mobility aids, history of falls and the International Classification of Functioning, Disability and Health (ICF) Multiple Sclerosis Brief Core set.40
Randomisation, blinding
Patients eligible will be provided with the study details. After obtaining written consent, patients will be randomly assigned using simple randomisation (Random Allocation Software for parallel group randomised trials41) and opaque sealed envelopes concealment. The allocation ratio is 1:1. Unblinding is permissible regarding participant safety, during the analysis, data interpretation or for regulatory compliance.
Retention and withdrawal, participation conditions
To promote adherence, the therapists will use their professional know-how and effective instruction to target each therapy to the current condition to increase adherence. Permitted concomitant care includes pharmacotherapy and other forms of supportive therapy—psychotherapy. Reasons for participant’s drop-out include discontinuation of the therapy for more than 2.5 weeks or five missed PTs, change in medication (by patient’s neurologist). The patient may step out at any time (Figure 1).
Participant timeline. Timeline showing key phases of the study chronologically. PT in VR (physical therapy in virtual reality, the experimental arm), PT (physical therapy, control arm), fMRI (functional MRI).
Intervention
Participants in both groups will undergo 2 months (8 weeks, 60 min two times per week) of ambulatory NFI physiotherapy, combining the key principles of the Proprioceptive Neuromuscular Facilitation (PNF) and the Motor Programme Activating Therapy (MPAT) methods. In MPAT, suitable afferent somatosensory stimuli are applied, affecting the associative brain cortex and initiating desirable motor programmes. The repetition of these activated programmes through a set of stimuli adjusts posture with anatomical centration of the joints under various conditions. This will lead to better support for postural stabilisation while seated, getting up, stepping forward and standing. This will help participants to automatically apply the acquired motor skills in their daily lives.42 PNF is a method used to learn effective movement patterns with high biomechanical efficiency based on repetitive stimulation of cooperating alpha-motoneurons and proprioceptors in muscles, tendons and joint capsules, using spiral and diagonal movements.43
Experimental group: NFI in VR
The VR environment is programmed as one of the facilitation stimuli to attract attention and motivate participants to perform movements in the practised seven therapeutic elements while guiding them into qualitatively correct patterns. Participants will be instructed to focus on the suggested trajectory of movement, visualised in immersive stereoscopic 3D. Each movement is further specified with a location point, rotation, orientation or directional hint arrow. Wrist, arm and trunk sensors continuously log positions and 3D orientations using HTC Vive Tracker 2.0 devices. The patient’s movements are projected in the VR for immediate feedback and displayed to the physiotherapist overseeing the therapy.
Each therapeutic session will consist of seven therapeutic elements: the therapeutic session will begin with the activation of the motor programme for sitting according to MPAT. This activated position, UL movements will be subsequently trained through a ball game, a counterclockwise spiral, and the activation of the qualitatively correct function of the upper extremities based on the PNF principles (diagonal I. II. and II. with flexion and rotation of trunk), followed by practice in standing up and sitting down (figure 2).
Virtual reality intervention description. (1) Activation of the motor programme for sitting according to MPAT. (2) A ball game. Participants collect 6–10 scattered balls starting with the right upper limb, followed by the left. (3) A counterclockwise spiral (4) PNF diagonals. In the activated sitting position, upper limb movements were performed using the PNF method in diagonal patterns and facilitated by appropriate afferent stimuli, including stretching, maximal resistance, manual contact, verbal cues, traction and compression. (5) Standing-up and sitting-down exercise. (6) The physiotherapist support in therapeutic sessions.
Each element will be repeated, with the number of repetitions gradually increasing based on the participant’s health condition—from seven repetitions in the first session to 12 by the end. The intensity is moderate. The physiotherapist actively engaged in the therapy, ensuring proper execution of tasks and providing facilitation, inhibition stimuli or support as needed. Emphasis was placed on movement quality, and if performed correctly, the therapist increased the activity’s difficulty using aids such as a TheraBand. Warm-up and cool-down exercises are not explicitly included in the protocol. Each therapeutic session will commence with feedback where participants will be asked about their postsession experience, including any perceived improvements or discomfort. In cases of musculoskeletal pain, the therapist will apply soft tissue techniques to address the issue. At the conclusion of each session, the therapist and participant will have time to review the recorded data, analyse the participant’s performance, highlight successful aspects and identify areas for correction. A more detailed description of the intervention is available in the online supplemental material 1.
Supplemental material
Active comparator group: NFI
The control group will receive therapy of equal duration, intensity and frequency (eight consecutive weeks, two times per week for 60 min), with the same seven therapeutic elements as outlined above, but without the use of VR. In the control group, the physiotherapist will also actively administer the NFI-based PT. The instructions include progressively increasing the number of repetitions based on individual performance (from seven to 12 per task).
Data collection and management
To promote the consistency of the data, instruments will be regularly calibrated. Both hands will be assessed. Separate Excel-based data sheets will be created for the baseline (E1) and the assessment after 2 months of therapy (E2; follow-up). All OMs will be examined within 1 week of the last therapy. The statistician and coordinator will conduct data vetting.
Outcome measures
(Figure 3)
Outcome measures. Outcome measures are evaluated at baseline and after the treatment. BBT, box and block test; CD-86, cluster of differentiation; DC, dendritic cells; ED-5Q-5L, EuroQol Questionnaire; fMRI, functional MRI; HGS, hand grip strength; HLA-DQ, human leucocyte antigen; lncRNA, long non-coding RNA; MoDC, monocyte-derived dendritic cell; MSIS-29, Multiple Sclerosis Impact Scale-29 item questionnaire; NHPT, nine hole peg test; 3D-FLAIR, three-dimensional fluid-attenuated inversion recovery; 3D MP-RAGE, magnetisation prepared-RApid gradient echo; 5STS, 5-times sit-to-stand test;
Primary OMs
Nine hole peg test (NHPT)
The NHPT is a standardised OM. It is a highly recommended OM.44 Its psychometric properties include excellent test-retest reliability45 and inter-rater and intrarater reliability46 with a minimal clinically important difference of 15,3%.47 The NHPT requires participants to repeatedly place and then remove nine pegs into nine holes, one at a time, as quickly as possible. Both ULs will be assessed with the NHPT (two trials per limb and averaged).
Box and block test (BBT)
The BBT is among the most used tests for assessing unilateral gross manual dexterity in MS48 and49 provides a valid assessment of UL function in MS with higher disability. Normative data are available for healthy adults.50 The number of cubes transferred with one hand from one compartment of a box to another of equal size within 60 s is counted. Two trials in each hand will be performed and averaged.
Hand grip strength (HGS)
Jamar Hand Dynamometer is a validated tool that measures isometric grip force and strength, measured three times for each position available (9 cm, 12 cm, 14,5 cm, 17 cm and 20 cm) in a standardised position (sitting with neutrally rotated shoulder, elbow flexed at 90°, forearm in a neutral position). Jamar Hydraulic Pinch Gauge measures finger pinch force—key pinch, pinch tripod and pinch tip-tip. In each position, three trials will be measured in each hand.
Tremor
An accelerometer device placed on the index finger records tremor for 1 min while holding one hand raised forward in a standing or sitting position. First, we will record with open eyes, and a second record will be measured with closed eyes. Four trials in total will be recorded, one for each position and one for each arm.
fMAX—the spectral characteristic of postural tremor measured by the 3-axis accelerometer and 3-axis gyroscope chip (motion tracking sensor MPU-6050)—a frequency for which the smoothed power spectral density is maximal.
Pf1-f2—the spectral characteristic of postural tremor, a power of the signal in a band from f1 to f2—lower value, lower tremor.
Five times sit to stand test (5STS)
The 5TSTS test is a highly reliable tool for assessing lower limbs strength, balance control and mobility.51 The time when the subject sits and stands repeatedly five times will be measured.
Secondary OM information
Multiple sclerosis impact scale (MSIS-29)
MSIS 29 is a highly recommended OM by MS EDGE and MSTF46 for use in clinical trials and also in an outpatient setting, due to the consistency of the results.52 A 29-item self-report measure with 20 items associated with a physical scale and nine items with a psychological scale. Summed responses are converted to a 0–100 scale, indicating the impact of the disease on daily function.53
EQ-5D-5L health questionnaire (EuroQol)
EuroQol is a descriptive system and an established and validated generic instrument for assessing health-related quality of life54 55 consisting of five dimensions.
Prespecified OMs
Kinematic analysis
The kinematic data from HCT Vive VR deliver suitable data quality for kinematic analysis56 to track the learning process and the subject’s understanding of the proposed trajectory in examination E1 versus E2, providing feedback on the accuracy (distance from the outlined trajectory, in metres) and duration of the movement. It is possible to measure the spatial distance from the suggested movement trajectory (scaled to fit each patient’s proportions) averaged for the exercise, as well as angular deviation from the suggested orientation of each tracked locus.
fMRI examination and analysis
The fMRI examination substudy will be performed in the Magnetic Resonance (MR) system Siemens Vida 3T, using a 64-channel Radiofrequency (RF) head coil. MR examination (E1 and E2) will include measurements of functional stimulated data related to motoric inputs, functional connectivity measurement (resting state measurement, individually assessed), structural images (three-dimensional SPACE (sampling perfection with application-optimized contrasts using different flip angle evolutions) FLAIR (fluid-attenuated inversion recovery) and three-dimensional magnetisation prepared-RApid gradient echo (3D MP-RAGE)) and structural connectivity, with 106 spatial directions and 3b-factors allowing the reconstruction not only of fractional anisotropy but also the diffusion kurtosis maps. All functional data will be coregistered to structural images acquired with isotropic high spatial resolution.
The protocol includes two stimulation intervals. Imaginary execution of screened exercise (10 s), followed by a resting state of 10 s and a 10 s unilateral hand active movement-driven stimulation interval. The fMRI measurement includes 320 volumes (each covering the brain with 72 slices) acquired using the echo-planar imaging (EPI) sequence with multiband slice excitation (four simultaneous slices, SMS4) and the following basic parameters: TE of 30 ms, TR of 1250 ms (temporal resolution), voxel size of 1.9×1.9×1.9 mm3 (spatial resolution). The resting state fMRI is measured with higher temporal resolution (TR of 650 ms) but slightly lower spatial resolution of 3×3×3 mm3.
The analysis will be performed using SPM 12 and CONN software. The fMRI data will be coregistered to individual structural 3D data with high spatial resolution (MP-RAGE with 1 mm3 isotropic spatial resolution). Statistical analysis on an individual level uses general linear model statistics and threshold for final statistical maps is p=0.05 with family-wise error (FWE) correction. When testing the fMRI examination protocol, four healthy volunteers repeatedly underwent examination to calculate the reproducibility and repeatability of searched effects (size and strength of brain motor activations).
Diffusion-weighted imaging data
MR scans will be performed according to an optimised protocol for diffusion-weighted imaging using a spin-echo EPI (SE-EPI) sequence; repetition time (TR)=4500 ms, echo time (TE)=92 ms, 69 axial slices, field of view (FOV)=200 mm, 106 diffusion directions and a voxel size of 2×2×2 mm³. A multishell diffusion scheme with b-values of 0 s/mm², 1000 s/mm² and 3000 s/mm² will be employed for baseline and follow-up scans. To compare longitudinal changes in white matter tracts differential tractography57 will be performed.
Immunological and epigenetic examination
Generation of MoDCs
A blood sample of 20 mL peripheral blood will be acquired in two 10 mL K3EDTA test tubes in examinations E1 and E2. PBMCs will be isolated by density gradient centrifugation using Ficoll-Paque media. Monocytic cell fraction will be separated by 2-hour cultivation in a 75 cm2 plastic culture flask in complete medium (CM) containing Roswell Park Memorial Institute (RPMI) 1640 medium, 10% fetal bovine serum, 1% antibiotic-antimycotic solution at 37°C in a 5% CO2 atmosphere. Adherent monocytes will be cultivated in CM supplemented with human granulocyte-macrophage colony‐stimulating factor and human IL‐4 for 6 days. MoDC maturation is induced by the addition of lipopolysaccharides to cells for 24 hours. Expression of the following CD cell markers will be analysed on leukocytes and MoDC: CD3, CD19, CD4, CD8, CD25, CD127, CD279, CD122, CD14, CD16, CD33, CD64, CD86, CD11c, HLA-DR and CD63 on a flow cytometer using antihuman monoclonal antibodies.
Analysis of lncRNA
Total RNA will be isolated from the second 10 mL K3EDTA test tube using the NucleoSpin RNA Midi blood Mini, according to the manual. Total RNA (1 µg) is converted into cDNA using standard protocol procedures and reagents (Thermo Fischer Scientific, Waltham, Massachusetts, USA).
Real-time quantitative PCR (qPCR)
The expression levels of preselected lncRNA (MALAT1, MEG3, H19, GAS5, PARTICLE and TINA) are determined by qPCR. For normalisation purposes and relative gene expression analysis, endogenous control genes are assessed (GAPDH, B2M) and measured in a StepOne eal-time PCR system instrument (VWR, Prague). All samples will be measured in triplets. The thermocycling conditions will be as follows: 50°C 2 min and, 95°C 10 min, followed by 40 cycles: 95°C 15 s and 60°C 1 min. Relative gene expression is quantified using the 2(-ΔΔCt) method.
Statistical analysis and software
The intervention and control groups will be compared in their characteristics and OMs. The normality of quantitative (continuous or discrete) outcomes will be explored using a quantile-quantile plot and Shapiro–Wilk’s test. The homogeneity of variances in the groups will be assessed by Levene’s test. The differences in means between the groups are evaluated using the independent samples t-test and within the groups using the paired t-test. The two-way mixed analysis of variance (ANOVA, between-subjects: group, within-subjects: time) will be performed to summarise and visualise the group and time effects on quantitative outcomes. If the assumptions of the above-mentioned parametric tests are not met, the data will be transformed using a Box–Cox normal transformation or non-parametric versions of tests will be applied (Wilcoxon rank-sum test, Wilcoxon signed-rank test, the two-way mixed ANOVA for aligned rank transformed data). The differences in the distribution of qualitative (categorical) outcomes between the groups will be evaluated using Pearson’s χ2 test or Fisher’s test, as appropriate, and within the groups using McNemar’s test. When multiple testing is done, procedures such as Bonferroni’s or Benjamini-Hochberg’s will be used to control for the so-called FWE.
We do not expect any significant differences in potential confounders (eg, the level of disability) between the groups at baseline. If they are, the statistical analysis will be adjusted (eg, using regression analysis, stratification or weighting) to control for selection bias. The attained level of significance (p value)<0.05 is considered statistically significant. R software is used for statistical data analysis.58
Harms
The PT intervention will be carried out by qualified specialists actively approaching all subjects concerning individual disabilities. In VR, the potential harms of immersion include cybersickness and/or spatial disorientation. The physiotherapists will attentively and actively approach to prevent these situations, and most exercises for ULs are carried out while sitting. The programme will be individualised to prevent patient exertion or harm. All adverse events must be reported to the principal investigator and relevant specialist and will be stored.
Strengths and limitations of the study
Specificity, precision and technique of the proposed movement are key features to the therapeutic effect combining flexibility regarding content, number of repetitions and movement performance with a motivating VR setting. The tested VR features seven unilateral UL tasks. To target bilateral engagement of the UL2 59 a VR software update is planned. Evaluation of VR contribution to the final effect is based on the intentional similarity of physiotherapy content in the real environment with VR PT. Possible sources of bias might occur from the simple randomisation process and include type, disease severity and MS stage, personal history and medication. Cognitive deficits regarding the participant’s ability to engage in the study will be evaluated by a treating neurologist. Participants will not be excluded for spasticity, as the therapeutic intent is to address it. However, its assessment could still be beneficial in future trials.
The duration of the therapeutic programme may be debatable concerning the potential to demonstrate clinical effects and plastic and adaptive changes in the CNS. However, based on our previous experience, our team believes this duration is sufficient and aligns with the lengths used in other studies.7 60–63
The proposed methods offer a complex evaluation of new VR using novel methods. There is no outright consensus on the fMRI protocol setting, design and interpretation found in the literature.64 Therefore, the proposed methodology could serve as a steppingstone to future comparisons of related RCTs with fMRI evaluation. The immunological and epigenetic substudy design serves as an exploration of molecular and cellular dynamics in response to PT, and it is a novelty within currently conducted PT studies in the multiple sclerosis patient population.
Results of this study have the potential to significantly contribute to the accessibility of innovative PT in outpatient care in PwMS, having a lifelong need for rehabilitation.
Ethics and dissemination
The multicentric ethics committee Faculty Hospital Kralovske Vinohrady and Medical Faculty of Charles University has approved the study under the code EK-VP/38/0/2021 (2 June 2021) and amendments (EK-VP/38/1/2021 on 4 April 2022). The ethics committee of the Institute for the Clinical and Experimental Medicine and Thomayer University Hospital have approved the study under No. 1983/21+4772/21 (G-21–02) (2 February 2021, and amendments (06958/22 (G21-26), 21 April 2022), based on submitted protocol, informed consent forms and participant education materials in Czech language (see English translation in the appendix). The estimated study completion is in January 2025. Study results will be disseminated to both the public and the scientific or medical community through publications in peer-reviewed journal articles and presentations at conferences, aiming to reach researchers and clinicians in the field. Anonymised data and research results may be shared on request for research purposes, such as meta-analyses or systematic reviews. Additionally, efforts will be made to communicate findings to the general public through collaborations with relevant organisations and the use of accessible briefs, ensuring broad accessibility and engagement.
Supplemental material
Ethics statements
Patient consent for publication
References
Footnotes
Deceased VBO deceased
Collaborators Not applicable.
Contributors KR, BM and LR—study conception, design and methods. LV and JF—virtual reality software development and optimisation. LR and AH—therapy content. IS, TP and JHl—site-specific establishment of PT. JT, II and JRy—the fMRI protocol for examination, reproducibility calculations and fMRI data analysis. JHa tremor measuring tool development and tremor data analysis, MC, MH, VBO’L, MP and IJ—lncRNA and dendritic cells conceptualisation and methods; JRe—data set and methods preparation and further data management (information upon request) and vetting. BM, KR, MH and JT writing—original draft. All authors contributed to the manuscript revision and approved the final version for submission. The deceased author, VBO’L, passed away prior to the manuscript submission but made extensive contributions to the process and fulfilled the other authorship criteria. KR is the guarantor of the study, taking full responsibility for the integrity of the work.
Funding The study was supported by the Grantová Agentura, Univerzita Karlova (Charles University Grant Agency) No 202322. The research was supported by Univerzita Karlova v Praze (Charles University), programme Cooperatio (Neuroscience and Medical Diagnostics and Basic Medical Sciences, the field ‘Medical Genetics’) No 260533/SVV/2024. The study was supported by SGS23/169/OHK3/3T/13 of the Czech Technical University in Prague. The study was supported (partly) by the long-term strategic development financing of the Institute of Computer Science (RVO:67985807) by Akademie Věd České Republiky (the Czech Academy of Sciences).
Competing interests None declared.
Patient and public involvement Patients and/or the public were not involved in the design, conduct, reporting or dissemination plans of this research.
Provenance and peer review Not commissioned; externally peer reviewed.
Author note The authors declare that all claims expressed in this article are solely those of the authors. The study sponsors provided financial support for the study's execution, including materials and operational costs. The VR software and tremor analysing tool were developed in cooperation with the principal investigator and other team members. Any other product that may be evaluated in this article or claim that may be made by its manufacturer is not guaranteed or endorsed by the publisher or manufacturer. A data monitoring committee was not established. Postclinical care is not contracted. All personal data are strictly confidential. Only pseudoanonymised data are analysed. Biological specimens are not stored. Access to the final trial dataset and statistical code will be restricted to the principal investigators and authorised research team members and will be made available upon reasonable request. Authorship for results will be granted if the coworker substantially contributes, drafts and revises manuscripts, is accountable and approves the final manuscript. The protocol SPIRIT 2013 and 2022 checklist is attached. Protocol V.V4/2024-10-24.
Supplemental material This content has been supplied by the author(s). It has not been vetted by BMJ Publishing Group Limited (BMJ) and may not have been peer-reviewed. Any opinions or recommendations discussed are solely those of the author(s) and are not endorsed by BMJ. BMJ disclaims all liability and responsibility arising from any reliance placed on the content. Where the content includes any translated material, BMJ does not warrant the accuracy and reliability of the translations (including but not limited to local regulations, clinical guidelines, terminology, drug names and drug dosages), and is not responsible for any error and/or omissions arising from translation and adaptation or otherwise.