The primary objectives are to investigate the effect of BFRp and BFRpE on cardiovascular safety in comatose patients in the ICU and to investigate the applicability of such protocols as early motor intervention methods in intensive care.
The secondary objective is to investigate the effects of this intervention on neuromuscular adaptations, physical function, and quality of life of ICU coma-surviving patients.
This study describes a protocol for a pilot study of a double-blind, randomized controlled clinical trial of applicability and intervention, with transverse temporality and a three-arm factorial intervention design in accordance with the Standard Protocol Items: Recommendations for Interventional Trials (SPIRIT) guidelines. The SPIRIT verification list can be found in the Additional File 1 and is represented in Fig. 1. The flowchart for the clinical trial is shown in Fig. 2.
The clinical trial will be carried out in the teaching and research ICU at the Irmandade da Santa Casa de Misericórdia de São Carlos, located in the city of São Carlos, São Paulo, Brazil. This ICU consists of 10 beds, and there is an average of 40 patients admitted monthly. Adult clinical and surgical patients with all types of diseases are treated in the ICU, and the average length of stay is 8 days.
Sample size rationale
A total of 30 participants were used for the sample size calculation of the clinical trial . Therefore, a sample of 39 patients will enable 10% sample loss for each experimental group (PM/control, BFRp, and BFRpE). Based on a recruitment average of 24 patients per month, 20% of non-consent and 30% of non-previewed death or immediate discharge, we aimed to recruit 7.2 patients per month. The recruitment in a single ICU will allow the study to be completed in a minimal period of 6 months.
The inclusion, non-inclusion, and exclusion criteria are shown in Table 1. All the criteria will be reevaluated before each training session, according to the clinical specificity of the criteria (Table 1). All non-included patients will be re-evaluated after 18 h of coma, except for patients whose hospitalization is longer than 18 h prior to ICU admission. In conclusion, all patients will be assisted by the ICU physiotherapy team. Additionally, the number of PM sessions or any type of physical therapy intervention in the peripheral muscle will be quantified to control the total volume of stimulus for each patient.
Initially, all hospitalized patients in the ICU will be assessed according to the inclusion criteria of the study. After screening, the legal representative will be interviewed and invited to be a participant in the study. Following consent, the legal representative will answer the functional performance of the basic activities of daily living through the Barthel Index  and the quality of life through the Medical Outcome Study 36 – Item Short Form Health Survey (SF-36) questionnaire [45, 46]. Following that, the patient will undergo other initial evaluations (T0) within a 23-h period after admission to the ICU or coma identification in the ICU hospitalization for the baseline demographic characterization, clinical, cardiovascular, and neuromuscular.
Initially, all participants will be pooled into a single group. Afterwards, patients will be classified into quartiles by muscle thickness (MT) of vastus lateralis (VL) and type of critical disease and assigned in a random and counterbalanced way into one of the three training protocols: PM/control, BFRp, or BFRpE using a maximally tolerated imbalance model . MT will be measured exclusively in the right leg. An unpaired t test will be used to ensure that there is no baseline difference between the groups. If significant between-group differences are detected, randomization will be repeated until a random distribution with no difference is achieved . The Berger-Exner test will be applied to verify selection bias .
Primary outcome measurements
Acute cardiovascular responses
The first training session (T1) will immediately succeed T0 and will have the monitoring of acute cardiovascular responses to training. Mean arterial pressure (MAP), central and peripheral systolic blood pressure (SBP [SBPc and SBPp]), and central and peripheral diastolic blood pressure (DBP [DBPc and DBPp]) will be measured by pulse wave analysis. Concomitantly, the heart rate (HR) and peripheral oxygen saturation (SpO2) should be recorded through a portable vital signal monitor (Dixtal, DX 2021, Amazonas, Brazil) every 2 min. The mean of 5 recordings at rest (10 min of rest), 18 recordings during exercise (~ 37 min of training), and the average of 5 post-exercise recordings (10 min) will be considered in the analysis. Electrocardiographic signals will be monitored, and, in case of adverse events, the session will be promptly interrupted. The training protocol will be applied to both legs during the session, simultaneously and alternately between the stimulated leg and the rest of the contralateral limb .
Cardiovascular safety and applicability of peripheral muscle training
To assess cardiovascular safety, HR values < 50 bpm or > 180 bpm, SBP < 80 mmHg or > 160 mmHg, DBP < 60 mmHg or > 80 mmHg, MAP < 60 mmHg or > 120 mmHg, cardiac or respiratory arrythmias, SpO2 < 88%, signals of respiratory discomfort [50, 51], or signals of deep venous thrombosis will be considered criteria to interrupt the protocol. Except for the occurrence of deep venous thrombosis, two consecutive training attempts will be performed after the first training interruption before withdrawing the patient from the study. It is suggested for two main reasons: to exclude the possibility of an occasional adverse event due to the variability of the clinical conditions, and to maintain the training frequency in case the patient continues in the study. In case of exclusion, the protocol will be considered unsafe for this patient. However, in case 15% or more unsafe conditions occur in the same training protocol, the protocol will be excluded from the study. To evaluate the applicability, the number of adverse events and intercurrence will be recorded, and if a total number of adverse events and/or intercurrence overcome 20% of the total training sessions, or there is deep venous thrombosis occurrence in at least 5% of the patients, protocols will be considered unsafe and impracticable. If cardiovascular variations are greater than 20% of the baseline, the protocol will be considered impractical.
Measurement of secondary outcomes
Neuromuscular adaptations of muscle mass and strength
Muscle mass and quality adaptations will be assessed daily prior to the first training session (~ 12 h from the last session) and 12 h after the last training session (T2) (until 12 h prior to the coma awakening), to the predicted death or ICU discharge. The muscle mass of the VL, rectus femoris (RF), and anterior tibial (AT) will be measured by MT and muscle quality, by echo intensity, both analyses obtained by ultrasound images. If the patient awakes from a coma and has the appropriate cognition to collaborate in active assessments, the global muscle strength will be evaluated by the Medical Research Council (MRC) score after 12 h of T2, in the ICU and at hospital discharge. Adequate cognition will be assessed by the absence of delirium after completing the Confusion Assessment Method for Intensive Care Units (CAM-ICU) score .
Physical function adaptations
Physical function will be assessed 12 h after T2 and in the ICU and at hospital discharge through the Physical Function in Intensive Care Test Scored (PFIT). Lastly, during home follow-up, functional motor performance of basic activities of daily living will be evaluated using the Barthel Index, 6 and 12 months after hospital discharge, by telephone.
Quality of life adaptations
Quality of life will be measured simultaneously with basic activities of daily living performance, using the SF-36 questionnaire, which is validated in Portuguese and Brazilian culture and is reliably applicable by telephone [53, 54].
To be considered a double-blind study, one of the authors will be assigned to create codes for the training-related adaptation data, in addition to being responsible for the randomization of patients. The evaluator, who evaluates the safety and training-related adaptations, will be blinded to the experimental groups. Each experimental group will be led by a different trainer who will also be blinded for the other training groups. Lastly, participants will be blinded by their coma, and in case the participant awakes from coma, information from the study will be provided, but not about the protocol they had previously undergone.
Peripheral muscle training
After T1, in view of the cardiovascular safety and applicability of PM, BFRp, and BFRpE, training sessions will be held twice a day, divided by 4 h . The starting training leg will be randomly chosen at T1 and, from that, alternated in every training session. Patients will undergo the experimental protocol if they present controlled temperature (between 36.5 and 37.5 °C), stable blood pressure (> 100 and < 150 of SBP and > 60 and < 100 of DBP), SpO2 > 90%, respiratory rate (RR) < 25 rpm, and HR > 60 and < 140 bpm .
Passive mobilization/control group protocol (PM)
The PM protocol will be modified from Barbalho et al. . The authors applied 3 sets of 15 mobilizations for flexion and extension of the hips, knees, and ankles. Movement was standardized in 2:2 using a metronome (2 s flexion and 2 extension), totaling 3 min of PM. To standardize PM with the BFR protocol, in every minute of PM, 1 min of rest will be allowed, totaling 5 min of PM for each set. Following that, a 3-min rest will be allowed, and the mobilization will be repeated in 5 sets. The session will last 37 min on average .
Passive mobilization associated with blood flow restriction protocol (BFRp)
A pressure cuff with 17.5 cm of width and 94 cm of length (JPJ, Sao Paulo, Brazil) with a pressure manometer will be placed on the quadriceps muscle, on the inguinal fold. Afterward, a vascular Doppler probe (DV-600; Martec, Ribeirao Preto, Brazil) will be placed in the posterior tibial artery. To determine the occlusion pressure, the cuff will be inflated until the auscultatory pulse stops . An external compression of 85% of the total vascular occlusion pressure will be applied for 5 min, followed by a 3-min rest (compression release) . The BFR protocol will be synchronized to the PM protocol. Compression will be repeated 5 times during the training session, and occlusion pressure will be re-evaluated in all sessions.
Passive mobilization associated with blood flow restriction and combined with neuromuscular electrical stimulation (BFRpE)
The procedures for BFRp will be adopted and synchronized with an electrical stimulation protocol. For the electrical stimulation electrode placement, the skin will be subjected to trichotomy, abrasion, and asepsis. Rectangular-shaped electrodes (50 × 50 mm) will be placed on the distal and proximal motor points of the VL (proximal and distal points located between the reference line of the antero-superior iliac spine and the superior lateral border of the patella and on the line between the apex of the great trochanter and the superior lateral border of the patella, respectively), RF (located on the reference line between the antero-superior iliac spine and the superior patella border), and AT (on the line between the apex of fibular head and the medial malleolus). In addition to the reference lines, the placement of the electrodes demonstrated in Santos et al.  and Dirks et al.  will be used to guide the location and electrostimulation of VL, RF, and TA. Additionally, this electrode placement will be marked with a semi-permanent pen to maintain the stimulus points throughout the study. A biphasic electro stimulator (Dualpex 071, Quark Medical, Piracicaba, Brazil) will provide symmetric pulses of 100 Hz, pulses lasting 400 μs, 5 s on (0.75 s rise, 3.5 s contraction and 0.75 s decay), and 10 s off, the intensity must be capable of causing visible contractions. The mean intensities used should be maintained from 29 to 33 mA and increased in approximately 3 min, when muscle contraction is no longer visible. The session will last 40 min, including 30 min of stimulation, a 5-min warm-up, and a 5-min cool-down, both with 5 Hz of frequency, pulse wave lasting 250 μs, and sub-maximal intensities (without muscle contraction) [59, 60]. The same researcher will apply both the BFRp and electrical stimulation.
Cardiovascular assessment by the pulse wave analysis
Pulse wave analysis will be performed using SphygmoCor equipment (SphygmoCor, AtCor Medical, Sydney, Australia). In a supine position, the patient will have a pneumatic clamp placed in the proximal portion of the left arm. Following that, additional data such as age, weight, and height will be provided to the system. After a 10-min rest, three measurements of systolic and diastolic arterial pressure will be performed in a 2-min interval  and accepted as long as the difference is less than 10% , or 5 mmHg  between the measurements. Afterward, the mean value of the three accepted measurements will be inserted into the system to obtain the MAP, SBPc, SBPp, DBPc, and DBPp [64,65,66,67]. In T0, the variables will be measured after 10 min of rest. In T1, it will be taken prior (10 min), during (23 and 37 min), and after (47 min) the training session. The average between minutes 23 and 37 (pause between sets) will be used as the exercise value.
Muscle mass and quality assessment
MT and echo intensity (MTecho) by ultrasonography
MT and MTecho will be measured by ultrasound images (Mysono U6 EX; Samsung Medison, Gangwon-do, South Korea). For body fluid homogenization, 15 min of rest in the supine position will be allowed, with the lower limbs neutrally positioned and sustained by a Velcro strip. A 7.5 MHz linear-array probe (LN5-12; Samsung Medison, Gangwon-do, South Korea), revested by surface gel, to promote acoustic coupling without dermic compression, will be placed longitudinally to the muscle fibers. The anatomic reference between two MT points of each muscle will be defined by the distal motor point sites used for the electrical stimulation. Additionally, for the VL, the anatomical reference will be the middle portion between the great trochanter and the lateral epicondyle . For RF, the reference will be set at 50% and at two-thirds of the distance from the anterior superior iliac spine and the superior patella border [51, 69]. Anatomical references for AT will be defined in one-fourth of the distance between the inferior portion of the patella and the lateral malleolus and one-third of the distance between the border of the fibula and the medial malleolus . Sequential images will be acquired with 6 cm of depth and, after digitalization, analyzed using the ImageJ software (National Institute of Health, USA). MT (cm) will be characterized as the perpendicular measurement between the superficial and deep aponeurosis . MTecho (A.U.) will be evaluated using greyscale grading, obtained by the histogram function of the software (0 = black and 256 = white) . Three consecutive images of the same anatomic point will be analyzed, and the mean will be adopted as the final value for MT and MTecho . MTecho will be normalized by MT (i.e., MTecho/MT, cm) to explain the effects of alterations on muscle size .
Muscle strength assessment
Global muscle strength by MRC score
Six muscle groups of the upper and lower limbs will be assessed bilaterally through the global strength (shoulder abductors, elbow flexors, wrist extensors, hip flexors, knee extensors, and feet dorsiflexors) by the MRC score [51, 74, 75]. Test familiarization will be performed through passive movements, and the tests will be applied in a fixed order. All muscle groups will be rated from 0 to 5, where 0 means absence of visible/palpable muscle contraction and 5 means movements performed against gravity and resistance . For the fragile and critical population, patients will be positioned in dorsal decubitus with the headboard angled at 45°. Then, arm abduction will be tested against gravity with the elbow in flexion. In case of a successful attempt, the strength will be rated from 4 to 5. Otherwise, patients will be repositioned in dorsal decubitus at 10°. to be rated from 0 to 3. Following that, the elbows, wrists, hips, knees, and ankles will be tested. In case any muscle group cannot be evaluated due to a central/peripheral nervous injury or orthopedic problems, the total sum will be calculated by extrapolating the identical contralateral muscle . The weakness acquired from the ICU will be identified from scores < 48  and classified as severe weakness if < 36 points .
Physical function assessment
Physical function by the PFIT scored
The PFIT scored will be applied to measure the physical function of the ICU patients. The score presents four items (assistance with sit to stand, cadence, strength of the shoulder flexors, and strength of the knee flexors), and for each item, the scoring varies from 0 to 3. The maximal score is 12, which indicates functional independence . The PFIT scored will be applied through the translated and validated version for the Brazilian population .
Physical function by the Barthel Index
The Barthel Index is widely used in Brazil [79, 80] and is applied in severely ill patients [81,82,83] and administered by telephone [84, 85]. It comprises 10 items weighted differently. Two items (bathing and personal care) can be graded in two alternatives (0 and 5 points), whereas 6 items (feeding, dressing, bowels [fecal and urinary continence], toilet use, stairs), in three alternatives (0, 5, and 10 points), and two items (transfers and mobility) present four alternatives (0, 5, 10, and 15 points). The final score is obtained by summing each item, varying from 0 (complete dependence) to 100 (independence) .
Quality of life assessment
Quality of life using the SF-36 questionnaire
Quality of life will be assessed using the SF-36 questionnaire, an instrument of good acceptance, reliability, and validity for ICU patients validated in Brazil and easily administered by telephone [45, 46, 53, 54]. It comprises 8 domains: physical function, role-physical, bodily pain, general health, vitality, social functioning, role-emotional, and mental health. Scores range from 0 to 100, where 0 corresponds to the worst general health status, and 100 corresponds to the best health condition [86,87,88]. The categorization and description of the methods and analyses are shown in Table 2.
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