Effect of Ultrasound-guided Lung Recruitment Maneuvers on Atelectasis in Lung-healthy Patients Undergoing Laparoscopic Gynecologic Surgery: a Randomized Controlled Trial

Atelectasis is a major cause of hypoxemia during general anesthesia and postoperative pulmonary complications (PPCs).Some previous reported that the combined use of lung recruitment procedures (LRMs) and positive end-expiratory pressure (PEEP) in mechanical ventilation mode contributes to the avoidance of PPCs in patients after general anesthesia, while others suggest that the use of LRMs makes patients more susceptible to hemodynamic disturbances and lung injury, and is of limited potential to decrease the incidence of PPCs. From this perspective, controversy exists as to whether LRMs should be routinely applied to surgical patients. More importantly, corresponding clinical studies are also lacking. Therefore, this trial was conducted with the aim of solving the above problem. Methods In clinical trial, patients undergoing laparoscopic gynecologic surgery with healthy lungs were randomized to the recruitment maneuvers group (RM group; 6 cm H 2 O PEEP and RMs) and the control group (C group; 6 cm H 2 O PEEP and no RMs). Lung ultrasound was performed on patients at ve separate time points. During mechanical ventilation, patients in the RM group received ultrasound-guided pulmonary resuscitation when atelectasis was detected, while the C group did not intervene. Lung ultrasound scores were used to evaluate the incidence and severity of atelectasis. for 24 no difference between the two groups. Date are presented as the mean±SD for continuous variables. PPCs, which included hypoxemia, pulmonary infection, atelectasis, acute respiratory failure, ventilator-associated lung injury, and neurogenic pulmonary edema. comparison between the two groups in per time point, with P<0.05 considered signicant.


Introduction
Mechanical ventilation is a common respiratory support method in general anesthesia, and atelectasis is one of the common complications of mechanical ventilation that can be responsible for the development of PPCs. It has been shown that the incidence of atelectasis after general anesthesia is up to 90% [1] and that it can happen at the induction of anesthesia and last up to two days postoperatively, which prolongs hospitalization and increases medical costs [2]. Atelectasis is the main cause of hypoxemia during general anesthesia. Patients undergoing gynecological laparoscopic surgery usually need to establish carbon dioxide pneumoperitoneum and Trendelenburg position, which may further increase the incidence of atelectasis and the possibility of hypoxemia, thereby promoting PPCs [3,4]. The reason may be that carbon dioxide pneumoperitoneum and Trendelenburg position employed in laparoscopic gynecologic will cause the abdominal contents to push the diaphragm more cephalad, resulting in aggravated lung collapse, decreased functional residual capacity and more prone to atelectasis during perioperative period [5][6][7].
Although PEEP alone can improve intraoperative oxygenation [8], studies have shown that the combination of LRMs and PEEP can better improve oxygenation and reduce the incidence of PPCs in patients [9,10]. Recent studies suggest that the combination of LRMs and PEEP is helpful to prevent the occurrence of PPCs after general anesthesia in adults [11][12][13]. However, whether LRMs can reduce PPCs remains controversial [14][15][16][17][18]. Therefore, it is still unclear whether LRMs should be routinely applied in patients with normal lung function and whether it is bene cial for patients need to be further explored.
CT is the gold standard for clinical diagnosis of anencephaly, but its application is somewhat limited by its relative disadvantages such as high radiation, high transport risk, costly and timeconsuming examinations [19,20]. Lung ultrasound is a simple bedside imaging method that features the advantages of being affordable, radiation-free, and portable compared to CT. More importantly, pulmonary ultrasound can reliably diagnose and monitor atelectasis [21][22][23]. Therefore, the application of lung ultrasound may be more bene cial to patients in situations where mobility is limited, such as postoperative, catastrophic, or epidemic situations. In conclusion, our study aimed to assess the impact of LRMs by pulmonary ultrasound on the incidence of atelectasis in the included patients and to provide references for the application of LRMs to surgical patients.

Study design
The prospective randomized controlled study was conducted from May 2020 to October 2020 and approved by the Ethics Committee of the A liated Hospital of North Sichuan Medical College (Ethical No. 2020ER079-1). All subjects signed an informed consent form and completed registration with the China Clinical Trials Center (Approval No. ChiCTR2000033529).

Study population
Inclusion criteria were patients with healthy lungs aged 18 to 65 years, with a body mass index (BMI) <35 kg/m 2 , American Society of Anesthesiologists (ASA) physical status I-II and undergoing gynecologic laparoscopic surgery. The pulmonary evaluation was performed by a specialized radiologist using X-rays and CT on the included subjects. Exclusion criteria were that patients with pulmonary disease, cardiac disease, neuromuscular disease and corresponding surgical history, as well as respiratory tract infections were also excluded. Withdrawal criteria were as follows: (1) patients with preoperative ultrasound suggestive of pulmonary atelectasis; (2) surgical conversion from laparoscopic to open; (3) serious postoperative complications such as severe subcutaneous emphysema and pneumothorax.

Randomisation and blinding
According to the computerized randomization software (www.randomization.com), patients were randomly divided into C group and RM group in the ratio of 1:1. Group assignments were concealed in sealed envelopes that were opened after the anesthesiologist administered general anesthesia to the patient. Except for the anesthesiologist performing the induction of anesthesia and pulmonary ultrasound, who knows the grouping details, neither the patient nor the pulmonary ultrasound evaluator is aware of the details.

Anesthesia and ventilation protocol
All patients received standard general anesthetic protocol. Including 5L·min −1 , 100% oxygen mask oxygen to nitrogen about 3 min, induction of 0.04 mg·kg −1 midazolam, 0.5µg·kg −1 sufentanil, 2 mg·kg −1 propofol, 0.6 mg·kg −1 rocuronium and the use of appropriate size of tracheal tube for intubation. The volumecontrolled mechanical ventilation mode was performed after intubation with a tidal volume of 8 ml·kg −1 , PEEP of 6 cmH 2 O and 0.4 inspired oxygen fraction (FIO 2 ).The initial respiratory rate was set at 12 breaths·min −1 with an inspiratory: expiratory ratio of 1:2. The ventilator was adjusted to maintain an endtidal carbon dioxide pressure (PETCO 2 ) at 35-45 mmHg. Anesthesiologists can adjust FIO 2 according to their experience when peripheral oxygen saturation <90%. Anesthesia was maintained by intravenous infusion of 0.1-0.3µg·kg −1 ·min −1 remifentanil and 4-12 mg·kg −1 ·h −1 propofol, inhalation of 1 %-3 % sevo urane. Bispectral index (BIS) was used to monitor the depth of anesthesia and maintain it at 40-60, timely supplement rocuronium to maintain adequate muscle relaxation. After spontaneous breathing recovery, neuromuscular blockade was reversed by neostigmine and glycopyrrolate. After extubation, the patient was sent to PACU, and oxygen was inhaled through the nose in PACU at a ow rate of 3 L·min The thorax was divided into left and right lungs, a total of 12 quadrants. Left and right lungs were divided into upper and lower zones. Each side was divided into anterior, lateral and posterior zones by anterior and posterior axillary line. In the anterior and lateral regions, the probe is placed upright to the costal space, and in the posterior regions the probe is placed parallel to the costal space. In order to quantitatively evaluate the severity of atelectasis, we used the modi ed lung ultrasound score from Monastesse et al [23].
Furthermore, the degree of the lung ultrasound score (LUS) was divided into four grades and scored between 0 and 3: (0) 0-2 B lines; (1) ≥3 B lines or 1 or multiple small subpleural consolidations separated by a normal pleural line; (2) multiple coalescent B lines or multiple small subpleural consolidations separated by a thickened or irregular pleural line; and (3) consolidation or small subpleural consolidation of >1×2cm in diameter. We de ned atelectasis to be signi cant if any region had the lung ultrasound score ≥2. The LUS was calculated by adding up the 12 individual quadrant scores, ranging from 0 to 36 points, with higher scores indicating chronic atelectasis. The primary endpoint was the incidence of intraoperative and postoperative pulmonary atelectasis and LUS score. The secondary endpoints were oxygen saturation, the incidence of intraoperative cardiovascular adverse reactions, PACU residence time, hospitalization time and PPCs.

Sample size estimation
We calculated the sample size using the data from previous studies. Yang et al.'s study shows that the frequency of atelectasis following a lung recruitment manoeuvre was 50%, compared with 95% in adults after laparoscopic colorectal surgery who did not receive a recruitment manoeuvre [17]. The sensitivity of lung ultrasound detection of atelectasis was 88% [24]. According to our preliminary experiment, the incidence of atelectasis was 81% in lung-healthy patients after gynecologic laparoscopic surgery, which was reduced to 40% by LRMs. Therefore, if we assume an alpha error of 0.05, power of 80% and allowing for a dropout rate of 10%, the required sample size would be 20 patients per group.

Statistical analysis
Anthropometric data and demographics were collected from the individual patient. Following the normality of data testing, the Mann-Whitney U test or t test was used for intergroup comparisons as applicable. Friedman test or paired t test was used for intra-group comparison. Chi-square test or Fisher's exact test was used for categorical variables. A two-sided P value less than 0.05 was considered signi cant unless Bonferroni adjustments were made. SPSS 25 and Graph Pad Prism 8 software were used for statistical analyses.

Results
From June to October 2020, 65 patients were included in this study. Among them, 23 patients were excluded for different reasons, as shown in Figure 1. The remaining 42 patients were randomly divided into the RM group and the C group. One patient in the RM group withdrew from the trial due to severe postoperative subcutaneous emphysema resulting in poorly visualized lung ultrasound. Finally, 21 patients in the C group and 20 patients in the RM group were included in the analysis (Figure 1). A total of 2460 images were collected. Representative lung ultrasound images at various periods are shown in Figure 2.
Baseline characteristics of patients included in the study are shown in Table 1. There were no signi cant differences. Intraoperation parameters of the enrolled patients were shown in Table 2, which were also similar between the two groups. Table 3, the incidence of atelectasis in both groups of patients at each time point. At T2, there was no statistical difference in the incidence of atelectasis between the two groups, and this state persisted until after the rst LRM. While after the second LRM, i.e., when observed at T4, the incidence of atelectasis in the RM group was lower than that in the C group,and this difference disappeared within 24 hours after operation (T5). Figure 3, lung ultrasound score of 12 lung regions from T1 to T5 in both groups. Only at T3, lung ultrasound scores (LUSs) were lower in the RM group compared with the C group. The difference in LUSs between the two groups was similar to the difference in the incidence of atelectasis, which also disappeared 24 hours after operation. Besides, there was no difference in PPCs between the two groups as shown in Table 2, and no side effects were found of recruitment manoeuvres in RM group.

As illustrated in
Lung ultrasound score of anterior, lateral and posterior regions in 2 groups from T3 to T5, as shown in Figure 4a-c. LUSs of posterior regions were higher than those of the other two regions from T3 to T5 in the same group, with no signi cant difference between the anterior and lateral regions. In addition, there were difference of LUSs between the two groups at T4 was mainly due to the difference of lung ultrasound score in posterior regions.

Discussion
In this prospective randomized controlled trial, we found that the combination of ultrasound-guided recruitment manoeuvres and PEEP could reduce the incidence of atelectasis in patients with PACU compared with PEEP alone, but this difference disappeared 24h after operation.
The incidence of atelectasis in the PACU was as high as 81% in the C group, whereas the incidence of atelectasis in the PACU was reduced to 40% in the RM group by ultrasound-guided recruitment manoeuvres prior to extubation. Although the incidence of atelectasis was different in PACU, there was no difference in oxygen saturation and residence time between the two groups. The possible reasons are as follows. First, patients were treated with nasal catheter oxygen inhalation after operation, and atelectasis can be effectively alleviated. Therefore, patients in the RM group and the C group can have no clinical symptoms such as hypoxia. Second, patients with healthy lung have mild postoperative atelectasis, and the remaining healthy lung units can meet the compensation of the body, so there are no clinical symptoms. Interestingly, 24 hours post-operation, we found that the difference in the incidence of atelectasis and lung ultrasound scores between the two groups no longer existed. Moreover, we observed that the hospital stay time of patients was not different. Therefore, we believe that the combination of recruitment manoeuvres and PEEP was not more bene cial in preventing PPCs after gynecologic laparoscopic surgery in lung-healthy adults undergoing general anaesthesia.
Although it is reported that there are few side effects of recruitment manoeuvres [25, 26] and it is not found in this experiment, recruitment manoeuvres still has the risk of conducting hemodynamic disorder and ventilator-induced lung injury [25]. Therefore, we believe that ultrasound-guided recruitment maneuvers are not necessary for gynecologic laparoscopic surgery patients with normal lungs.
Some studies have shown that alveolar collapse and atelectasis can be formed within 5 min after general anesthesia induction and can persist postoperatively [2,27]. In present study, we found that in patients undergoing laparoscopic gynecological surgery, atelectasis can be formed after intubation and still not completely disappeared 24 hours postoperatively, which is in agreement with the results of the relevant studies [2,27]. is an important in uencing factor for atelectasis, and related literature reports that the incidence of atelectasis during general anesthesia for laparoscopic surgery is positively correlated with FIO 2 ranged from 0.4 to 1. We administered pure oxygen during induction, which may be responsible for the development of alveolar collapse in patients several minutes after induction [28,29].
Notably, there was no signi cant difference in the incidence of atelectasis between the two groups during operation. It can be seen that the combined application of ultrasound-guided lung recruitment maneuvers and PEEP after intubation had no signi cant effect on the progress of intraoperative atelectasis compared with the simple application of PEEP. Possible reasons for this are that intraoperative factors causing atelectasis continually take effect and that the short-term advantage from LRMs is not sustained, or because PEEP is relatively small and does not preserve alveolar opening. In our study, atelectasis mostly appears in the posterior regions of the lung (the patient's dorsum in a supine state), which is consistent with the view that atelectasis is mainly concentrated in the gravity-dependent area [23].
Before extubation, all sbujects in the RM group underwent the second LRM under ultrasound guidance until the atelectasis disappeared completely. When the lung ultrasound was reexamined in PACU after extubation, it was found that atelectasis was still present in 40% of the patients, and the alveolar collapsed again in a short time. The reason may be that the patients were in the PACU in the supine position with cranial displacement of the diaphragm, or other reasons such as postoperative extubation, obstruction by secretions, insu cient metabolism of anesthetic drugs, weak respiratory motility, and severe pain,etc [30].
Studies have shown that ultrasound-guided LRMs is more effective in diminishing atelectasis incidence in children than conventional methods [31], so we employed ultrasound-guided LRMs to minimize its undesirable effects. This study also suggests that it is feasible to monitor the changes of lung aeration in perioperative by lung ultrasound, and it can continuously and dynamically track the changes of aeration loss, which can be more widely applicable in the study of mechanical ventilation.
Of course, this study has some limitations. 1. We administered pure oxygen from induction of anesthesia to the end of intubation in order to improve anesthetic safety, rather than optimal FIO 2 . The increase in FIO 2 will contribute to the expansion of atelectasis area [32], possibly due to accelerated absorption of alveolar gas, resulting in absorptive atelectasis. 2. Our study included patients with healthy lungs and short surgery, and the incidence of PPCs was expected to be lower than after major surgery. Therefore, the preventive effect of ultrasound-guided LRMs on postoperative atelectasis may not be signi cant. It is necessary to further study the patients with high risk of PPCs.

Conclusions
In conclusion, our data indicate that the combination of LRMs and PEEP can reduce the incidence of atelectasis 15 min after arrival in PACU, but this advantage disappeared within 24 hours after operation, and it cannot improve PPCs in lung-healthy adults. Consequently, we do not recommend routine recruitment maneuvers for patients with healthy lungs undergoing gynecologic laparoscopic surgery.

Availability of data and materials
The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.    Figure 1 Flow diagram of patient screening and enrollment.  Lung ultrasound score of 12 lung regions from T1 to T5 in both groups. The box, whiskers and bold line across the box represent interquartile range, range and the median value, respectively. ns, no signi cance; ***, P 0.05.

Figure 4
Lung ultrasound score of anterior, lateral and posterior regions in 2 groups from T3 to T5. The box, whiskers and bold line across the box represent interquartile range, range and the median value, respectively. *P < 0.05, posterior region vs. anterior region in the same group; #P < 0.05, posterior region vs. lateral region in the same group; †P < 0.05, RM group vs. C group.