Changes in Quantitative Muscle Ultrasound for Diagnosis of Intensive Care Unit-Acquired Weakness in Critically Ill Patients

Background: A common occurrence in severe illness is muscle wasting, which is dened as intensive care unit acquired weakness (ICU-AW) and characterized by accid tetraparesis with areexia or hyporeexia. Many studies revealed the tendency of changes in quantitative muscle ultrasound parameters in critical illnesses; however, the relation between those changes in muscle parameters and intensive care unit acquired weakness was unknown. Objectives: Using the Medical Research Council Criteria, test the accuracy in diagnosis of changes in quantitative muscle ultrasound for diagnosing intensive care unit acquired weakness. Methods: Patients who were conscious and positively responded to verbal commands with facial muscles were subjected to quantitative muscle ultrasonography including measuring thickness and cross-sectional area of biceps brachii (BB) muscle, vastus intermedius (VI) muscle, and rectus femoris (RF) muscles for 4 times in the next 10 days after assessing their muscle strength using the Medical Research Council score. Assessment of the diagnostic accuracy of changes in cross-sectional area and thickness of different muscle groups was made using area under the curve of the receiver operating characteristic curve (ROC–AUC). We also calculated specicity, sensitivity, negative predictive value, positive predictive value, and diagnostic accuracy of 15% threshold for reduction of thickness and 12% threshold for reduction of cross-sectional area. Results: 37 patients underwent muscle ultrasonography for 4 times and 24 were found to have ICW-AW. ROC–AUC of changes in muscle parameters were acceptable and ranged from 0.647 to 0.727. The changes in the thickness of vastus intermedius muscle on both sides while the change in thickness and cross-sectional area of rectus femoris muscle on the right side showed good diagnostic accuracy and ranged from 75.7% to 78.4%. Conclusions: Ultrasonography of the changes in muscle parameters with good diagnostic accuracy is an alternative approach for making a diagnosis of intensive care unit acquired weakness. Particularly, changes in the thickness of vastus intermedius muscle on both sides and changes in thickness and cross-sectional area of rectus femoris muscle on the right side have diagnostic potential.

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)).

Study procedures
After enrollment, fully conscious patients were assessed for muscle strength by using the MRC score [22]. For the patients mechanically ventilated with sedative, if the RASS (Richmond Agitation Sedation Scale) fell anywhere between −1 and 1 [23] and they showed a positive reaction to 5 verbal commands with facial muscles, we considered them feasible for muscle strength assessment [11]. Twelve muscle groups need to be tested for the calculation of MRC score including elbow exion, wrist extension, shoulder abduction in upper limbs, and dorsi exion of the foot, hip exion 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 [12]. The MRC score is extensively utilized for diagnosing the ICU-AW and its good interobserver reliability in critical settings was con rmed in the former study [11].
After strength assessment, other researchers who were trained and blinded for the muscle strength results started to measure the muscle parameters immediately for the rst time by using a Philips ultrasound machine (IU22, USA) and a linear transducer or probe (frequency: 10-13MHz) which enabled acquiring images of super cial structures in high resolution [24]. 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 [25]. 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 [26], 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 [25]. When performing ultrasonography, the pressure to the skin was kept minimal, and adequate coupling agents were used for obtaining the images [3]. 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 nal value. The whole muscle ultrasound procedure was repeated on day 4, day 7, and day 10 to know the changes of these muscle parameters.

Sample size
According to the equation of diagnostic experiment and our preliminary data [27], 33 examined subjects were found to be su cient to detect discrimination e ciency in order to provide 90% power having a value of 0.05 for a two-sided level. We expected the sensitivity and speci city 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.

Statistical analysis
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% con dence 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 = (CSA day10 -CSA day1 ) / CSA day1 or ΔTH = (TH day10 -TH day1 ) / TH day1 . Based on ROC curve analysis, speci city, sensitivity, positive and negative predictive values (PPV, NPV) for muscle ΔCSA and ΔTH were calculated. A signi cant two-level p-value taken for all analyses was <0.05. SPSS version 19 was used to carry out all statistical analyses.  Table 1 lists patient characteristics. Whether or not patients suffered from ICU-AW, the groups all presented a descending trend of both TH and CSA bilaterally. In the upper limbs, only the changes of CSA in BB muscle on the right side showed a statistical difference in ICU-AW patients and non-ICU-AW individuals. Moreover, ICU-AW patients had an obviously greater degree of declination in CSA of RF muscle bilaterally, and a remarkable reduction of TH in VI muscle as well (Table 2). ΔTH of BB muscle on both sides had higher ROC-AUC than CSA, but ΔCSA of RF muscle had higher ROC-AUC than TH, especially on the right side. ΔTH and ΔCSA of RF muscle on the right side showed the highest ROC-AUC which was 0.840 and 0.888 respectively. Additionally, ΔTH of VI muscle also presented good diagnostic value for the diagnosis of ICU-AW. (Table 3, Figure 2 &3). Further, using speci c thresholds (15% for ΔTH of BB, RF and VI muscle, 12% for ΔCSA of BB and RF muscle) in term of Youden Index of ΔTH and ΔCSA that were calculated based on the ROC curve, sensitivity, speci city, PPV, NPV, and accuracy were con rmed and presented in Table 4. Diagnostic accuracy of ΔTH and ΔCSA of RF muscle on the right side and ΔTH of VI muscle on both sides was high and ranged from 75.7% to 78.4%.

Discussion
The monitoring and diagnosis of ICU-AW are challenging in ventilated and sedated ICU patients because clinical examination of strength particularly needs cooperation from patients, which is an obvious disadvantage in evaluation [19,28]. In contrast, muscle ultrasound is a non-invasive, painless, and commonly used bedside technique that is entirely independent of patient cooperation. It is hence of great signi cance for assessing and monitoring muscular waste in critically ill patients [13,21]. The present study con rmed that patients with ICU-AW had a signi cant reduction of muscle TH and CSA than those of patients without ICU-AW, especially in the lower extremities. Moreover, for a 15% threshold for reduction of TH and a 12% threshold for reduction of CSA, muscles in lower extremities showed good diagnostic accuracy of the diagnosis of ICU-AW, particularly on the right side.
In this study, of all evaluated patients, 64.9% were found to have ICU-AW. Whether or not patients had ICU-AW, all patients presented the descending trend of both TH and CSA bilaterally. Moreover, patients with ICU-AW had an obviously greater degree of declination in CSA of RF muscle bilaterally, and a remarkable reduction of TH in VI muscle of both sides as well. Turton et al. carried out a study on 22 ICU patients who were mechanically ventilated and performed an ultrasonographic assessment of the exor compartment of the elbow joint, the vastus lateral muscle and the medial head of the gastrocnemius muscle on admission and 10 days later. The results showed that the loss of muscle mass mainly occurred in the lower extremities and there was no change in the size of the exor compartment of the elbow joint. Such data helps to justify the argument regarding investigating the lower extremities further as peripheral muscles, in patients with critical illnesses have more chances to undergo early disuse atrophy [29]. In particular, a 3-week follow-up study employed ultrasonography to evaluate RF muscle in terms of the morphological changes and found severe muscle mass loss in CSA and muscle diameter experienced by all the ICU trauma patients. By day 20, approximately 45% of RF muscle mass was lost [30]. Consequently, in comparison to upper limbs, lower limbs muscles experience earlier and greater atrophy. The potential reason was given in an earlier study that assessed CSA of RF muscle and protein/DNA ratio over time. The results showed that during the rst week virtually all cases decreased in muscle mass. Lower limb muscle atrophy is considered to be the result of net catabolism due to the decrease of muscle protein synthesis and the simultaneous increase of protein decomposition relative to protein synthesis [15].
Few previous studies tested muscle ultrasonography for the diagnosis of ICU-AW or prediction of prognosis or similar symptoms. One study diagnosed skeletal muscle loss by measuring the CSA of RF muscle using ultrasound and compared it with frailty to predict the prognosis of critically ill patients. The outcomes of the study clearly show that the prediction value of adverse discharge tendency by bedside ultrasound in the diagnosis of skeletal myopenia was consistent with frailty [31]. Moreover, a prospective observational study found that larger the CSA of RF muscle on the day of admission, lower the occurrence of the muscle ber necrosis and muscle waste of RF muscle [18]. In addition, Greening et al. demonstrated that an independent risk factor for unscheduled readmission or death is a smaller quadriceps muscle size measured by ultrasound [32]. These studies showed the potential diagnostic value for ICU-AW diagnosis. However, these studies did not test the possibility of discrimination value of the muscle parameters by ultrasound between patients with and without ICU-AW. Witteveen's study performed ultrasonographic TH of tibialis anterior muscle, BB muscle, exor carpi radialis muscle, and RF muscle thus nding that for these muscles, the diagnostic accuracy of muscle parameters was rather low with ROC-AUC ranging from 51.3 to 68.0% [21]. Nevertheless, the study failed to observe the changes in muscle parameters over time, and CSA which is considered as a crucial property for contraction and strength of muscle was not involved in the study [13]. To the best of our knowledge, the present study is the rst of its kind testing the usefulness of changes in muscle parameters by using ultrasound to diagnose ICU-AW. According to results, the changes in quantitative muscle ultrasound had good performance when analyzed on MRC criteria for the diagnosis of ICU-AW, which endorses the use of muscle ultrasound as a surrogate tool for ICU-AW diagnosis. We also con rmed that the best cutoff value of reduction in muscle parameters for diagnosing ICU-AW using ultrasound is more than 15% for TH in VI muscle and more than 12% for CSA in RF muscle of the right side.
The ndings of the present study con rmed that muscle ultrasound is an alternative approach for earlier recognition of ICU-AW, especially the unconscious patients who are not suitable for strength assessment. However, some limitations of this study deserve the necessary commentary. First, due to the limited availability of biopsy or electroneuromyography in the ICU, we could not classify patients with ICU-AW into the three subcategories (critical illness neuromyopathy (CINM), critical illness myopathy (CIM) and critical illness polyneuropathy (CIP)). Second, we did not observe other ultrasonographic characteristics of muscle in recognition of ICU-AW, for instance, pennation angle and echo intensity which may have better diagnostic value. Last but not the least, it was impossible for ultrasound examiners to be completely blind to the MRC score because the absence or presence of spontaneous movements gave the impression of muscle strength. Therefore, in order to improve the accuracy of muscle measurement, all CSA and TH were measured three times in a row, and the average value is calculated as the nal value to minimize the performer's deviation.

Conclusion
Ultrasound measurement of muscles is an alternative tool for management and recognition of ICU-AW. Changes in TH and CSA of RF muscle on the right side and the changes in TH of VI muscle on both sides had good diagnostic accuracy for diagnosis of ICU-AW.

Declarations
Ethics approval and consent to participate The study was reviewed and approved by the Ethics Committee of the Rui Jin hospital (Reference Number 2018-90) in August 2018.
Written informed consent was obtained from all participants or their immediate relatives.

Competing interests
The data generated from this study will be made available on request to the corresponding authors. Authors' contributions WQZ contributed to design and study protocol preparation, and prepared the manuscript's rst draft, and was responsible for set-up and day-to-day conduct of study at the department, and obtained funding from School of Medicine, Shanghai Jiao Tong University. JW contributed to design and study protocol preparation, and trained YJZ, QZ, QL in ultrasound assessment. QYG contributed to design and study protocol preparation, and facilitated set-up of study at site, and was involved in manuscript preparation. YTG prepared the Statistical Analysis Plan, was involved in manuscript preparation and was involved in the statistical analysis of the data. YJZ, XYG, XJS, QZ were responsible for day-to-day conduct of study at the department, and assisted in authoring the manuscript. All authors read and approved the nal manuscript.
Flowchart representing screening and inclusion of patients.  ROC curves for ΔCSA of muscles