Population and Design
The present study designed to be carried out at a single center, a general ICU in Shanghai, China from June 2019 to May 2020. The study was duly approved by the Ethics Committee of the hospital concerned and informed consent to participate (either directly or through an appropriate surrogate) was taken from all patients. Patients were aged ≥ 18 years, their anticipated ICU stay was at least 2 days, and was eligible for screening after being evaluated daily for awakening and reaction to simple verbal command. Exclusion patients comprised individuals with prior diagnosed diseases characterized by generalized or regional weakness or with any diagnosis at the time of admission making patients abnormal muscle strength and unable to follow commands (e.g., cardiac arrest, stroke, spinal injury, traumatic brain injury or intracerebral infection), or delirium or dementia during the ICU stay. Additionally, the patients experiencing edema of upper and lower limbs and patients who did not have arms or legs for muscle strength testing or ultrasound or had wounds, fractures, lesions, burns, or bleeding at the measurement points were excluded as well.
Clinical data collection
Baseline data was collected after ICU admission and included age, sex, Body Mass Index (BMI), admission diagnosis, Sequential Organ Failure Assessment (SOFA) score, Acute Physiology and Chronic Health Evaluation II (APACHE II) score, risk factors for polyneuropathy or myopathy (restraints, surgery, nutritional supports, mechanical ventilation, glucose peak concentration, glucocorticoid, use of sedative and analgesic) and comorbidities (cardiac dysfunction, respiratory failure, liver dysfunction, acute kidney injury, hypertension, diabetes mellitus and Multiple Organ Dysfunction Syndrome (MODS)).
After enrollment, fully conscious patients were assessed for muscle strength by using the MRC score . For the patients mechanically ventilated with sedative, if the RASS (Richmond Agitation Sedation Scale) fell anywhere between −1 and 1  and they showed a positive reaction to 5 verbal commands with facial muscles, we considered them feasible for muscle strength assessment . Twelve muscle groups need to be tested for the calculation of MRC score including elbow flexion, wrist extension, shoulder abduction in upper limbs, and dorsiflexion of the foot, hip flexion knee extension in lower extremities. The function of each muscle group ranged from 0 to 5 and a mean MRC score <4 or the total MRC score <48 was categorized as the occurrence of ICU-AW in terms of the international consensus statement . The MRC score is extensively utilized for diagnosing the ICU-AW and its good interobserver reliability in critical settings was confirmed in the former study .
After strength assessment, other researchers who were trained and blinded for the muscle strength results started to measure the muscle parameters immediately for the first time by using a Philips ultrasound machine (IU22, USA) and a linear transducer or probe (frequency: 10–13MHz) which enabled acquiring images of superficial structures in high resolution . Before performing, the patient must be in a supine position with extended elbows, wrists, knees and relaxed muscles, meanwhile palms and toes of patients were facing or pointing to the ceiling . The muscle parameters for ultrasonography including CSA and TH of biceps brachii (BB) muscle, vastus intermedius (VI) muscle, and rectus femoris (RF) muscles. All the muscles were measured bilaterally and scanned in the transversal (cross-sectional) image. The transducer was oriented transversally in relation to the longitudinal axis of the arm or thigh for obtaining a cross-sectional image, thus creating a right angle to the skin surface. Landmarks for ultrasound image acquisition were at standardized anatomical points, including the midpoint between supraglenoid tubercle and radial tuberosity for BB muscle , the second third of the distance between the anterior inferior iliac spine (AIIS) and the midpoint of the proximal border of the patella for RF muscle, and the midline of the same distance as RF muscle for VI muscle . When performing ultrasonography, the pressure to the skin was kept minimal, and adequate coupling agents were used for obtaining the images . In order to enhance the accuracy of the measurement of target muscles, all the CSA and TH were measured three times continuously and an average was calculated as the final value. The whole muscle ultrasound procedure was repeated on day 4, day 7, and day 10 to know the changes of these muscle parameters.
According to the equation of diagnostic experiment and our preliminary data , 33 examined subjects were found to be sufficient to detect discrimination efficiency in order to provide 90% power having a value of 0.05 for a two-sided level. We expected the sensitivity and specificity of muscle ultrasonography to be 0.8. Based on preliminary data, ICU-AW was 50% prevalent in our critically ill patients. Thus, it was decided upon enrolling 36 subjects, considering the loss of 10% of the sample.
Kolmogorov-Smirnov’s normality test was employed for evaluating continuous variables’ distribution. Data acquired from continuous variables with a normal distribution are expressed either as mean and standard deviation or as the interquartile range (IQR) or median in case if they have a non-normal distribution. Mann-Whitney test, Student t-test, exact Fisher test, and chi-squared test were employed to assess the differences among patients with and without ICU-AW diagnosis according to the distribution and type of the variable.
The discriminative power of changes of muscle ultrasound over time was examined with a 95% confidence interval (CI) using ROC-AUC. The change of CSA and TH are represented by ΔCSA and ΔTH respectively, and was calculated using the formula: ΔCSA = (CSAday10 – CSAday1) / CSAday1 or ΔTH = (THday10 – THday1) / THday1. Based on ROC curve analysis, specificity, sensitivity, positive and negative predictive values (PPV, NPV) for muscle ΔCSA and ΔTH were calculated. A significant two-level p-value taken for all analyses was <0.05. SPSS version 19 was used to carry out all statistical analyses.