New Methods for Optimization of Mechanical Ventilation

Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine 1102

New methods for optimization of mechanical ventilation

PETER KOSTIC

ACTA UNIVERSITATIS UPSALIENSIS ISSN 1651-6206 ISBN 978-91-554-9238-0 UPPSALA urn:nbn:se:uu:diva-249172 2015 Dissertation presented at Uppsala University to be publicly examined in Enghoffsalen, Entrance 50, Akademiska sjukhuset, Uppsala, Wednesday, 3 June 2015 at 09:15 for the degree of Doctor of Philosophy (Faculty of Medicine). The examination will be conducted in Swedish. Faculty examiner: Johan Petersson (Karolinska institutet, Institutionen för fysiologi och farmakologi, Anestesiologi och intensivvård).

Abstract Kostic, P. 2015. New methods for optimization of mechanical ventilation. Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine 1102. 62 pp. Uppsala: Acta Universitatis Upsaliensis. ISBN 978-91-554-9238-0.

Mechanical ventilation saves lives, but it is an intervention fraught with the potential for serious complications. Prevention of these complications has become the focus of research and critical care in the last twenty years. This thesis presents the first use, or the application under new conditions, of three technologies that could contribute to optimization of mechanical ventilation. Optoelectronic plethysmography was used in Papers I and II for continuous assessment of changes in chest wall volume, configuration, and motion in the perioperative period. A forced oscillation technique (FOT) was used in Paper III to evaluate a novel positive end-expiratory pressure (PEEP) optimization strategy. Finally, in Paper IV, FOT in conjunction with an optical sensor based on a self-mixing laser interferometer (LIR) was used to study the oscillatory mechanics of the respiratory system and to measure the chest wall displacement. In Paper I, propofol anesthesia decreased end-expiratory chest wall volume (VeeCW) during induction, with a more pronounced effect on the abdominal compartment than on the rib cage. The main novel findings were an increased relative contribution of the rib cage to ventilation after induction of anesthesia, and the fact that the rib cage initiates post-apneic ventilation. In Paper II, a combination of recruitment maneuvers, PEEP, and reduced fraction of inspired oxygen, was found to preserve lung volume during and after anesthesia. Furthermore, the decrease in VeeCW during emergence from anesthesia, associated with activation of the expiratory muscles, suggested that active expiration may contribute to decreased functional residual capacity, during emergence from anesthesia. In the lavage model of lung injury studied in Paper III, a PEEP optimization strategy based on maximizing oscillatory reactance measured by FOT resulted in improved lung mechanics, increased oxygenation, and reduced histopathologic evidence of ventilator-induced lung injury. Paper IV showed that it is possible to apply both FOT and LIR simultaneously in various conditions ranging from awake quiet breathing to general anesthesia with controlled mechanical ventilation. In the case of LIR, an impedance map representing different regions of the chest wall showed reproducible changes during the different stages that suggested a high sensitivity of the LIR-based measurements.

Keywords: mechanical ventilation, optoelectronic plethysmography, forced oscillation technique, laser interferometry

Peter Kostic, Department of Surgical Sciences, Anaesthesiology and Intensive Care, Akademiska sjukhuset, Uppsala University, SE-75185 Uppsala, Sweden.

ISSN 1651-6206 ISBN 978-91-554-9238-0 urn:nbn:se:uu:diva-249172 (http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-249172)

To my family

List of Papers

This thesis is based on the following papers, which are referred to in the text by their Roman numerals.

I Aliverti, A., Kostic, P., Lo Mauro, A., Andersson-Olerud, M., Quaranta, M., Pedotti, A., Hedenstierna, G., Frykholm, P. (2011) Effects of propofol anaesthesia on thoraco-abdominal volume variation during spontaneous breathing and mechanical ventilation. Acta Anaesthesiol Scand, 55(5):588-96 II Kostic, P., Lo Mauro, A., Larsson, A., Pedotti, A., Hedenstier- na, G., Frykholm, P., Aliverti, A. (2014) Active expiration may contribute to the reduction in end-expiratory volume during emergence from anesthesia and in the immediate post-operative period. In manuscript III Kostic, P., Zannin, E., Andersson-Olerud, M., Pompilio, P.P., Hedenstierna, G., Pedotti, A., Larsson, A., Frykholm, P., Del- laca, R. (2011) Positive end-expiratory pressure optimization with forced oscillation technique reduces ventilator induced lung injury: a controlled experimental study in pigs with saline lavage injury. Crit Care, 15(3):R126 IV Kostic, P., Milesi, I., Zannin, E., Larsson, A., Frykholm, P., Dellaca, R.L. (2015) The application of forced oscillation technique and self-mixing laser interferometer during anesthesia and mechanical ventilation. In manuscript

Reprints were made with permission from the respective publishers.

Contents

Introduction ...... 11 Background ...... 13 Mechanical ventilation and ventilator-induced lung injury ...... 13 Protective lung ventilation ...... 13 Forced oscillation technique ...... 15 Experimental setup ...... 16 Laser interferometry ...... 17 Experimental setup ...... 17 Effects of anesthesia on respiratory function and anesthesia-induced atelectasis ...... 18 Anesthesia and postoperative pulmonary complications ...... 19 Lung-protective ventilation strategy during anesthesia and the perioperative period ...... 20 Optoelectronic plethysmography ...... 22 Aims ...... 24 Materials and methods ...... 25 Papers I and II ...... 25 Patients ...... 25 Anesthesia ...... 25 Protocol ...... 26 Data analysis ...... 28 Paper III ...... 29 Animal model and preparation ...... 29 Protocol ...... 29 Data analysis ...... 30 Paper IV ...... 30 Patients ...... 31 Anesthesia ...... 31 Protocol ...... 31 Data analysis ...... 31 Statistical analysis (Papers I-IV) ...... 32

Results ...... 33 Paper I ...... 33 Paper II ...... 36 Paper III ...... 39 Paper IV ...... 42 Discussion ...... 45 Main findings ...... 45 Papers I and II ...... 45 Paper III ...... 47 Paper IV ...... 49 Conclusions ...... 50 Limitations ...... 51 Future perspectives ...... 52 Acknowledgements ...... 53 References ...... 55

Abbreviations

ABW actual body weight ALI acute lung injury ANOVA analysis of variance APL adjustable pressure limiting (valve) ARDS acute respiratory distress syndrome ASA American Society of Anesthesiology BMI body mass index Cdyn dynamic compliance CPAP continuous positive airway pressure Crs respiratory system compliance CT computed tomography Cx5 oscillatory compliance derived from Xrs Ers respiratory system elastance FIO2 fraction of inspired oxygen FOT forced oscillation technique FRC functional residual capacity Irs inertance i.v. intravenous/ly LIR laser interferometer LMA laryngeal mask airway MAP mean arterial pressure MPAP mean pulmonary arterial pressure OEP optoelectronic plethysmography PaCO2 partial pressure of arterial carbon dioxide PaO2 partial pressure of arterial oxygen PBW predicted body weight PCV pressure controlled ventilation PEEP positive end-expiratory pressure PEEPol open lung PEEP PPC postoperative pulmonary complication Pplat plateau pressure PSV pressure support ventilation

Rdyn dynamic resistance RM recruitment maneuver RMS root mean square of EMG signal Rrs respiratory system resistance SB spontaneous breathing SD standard deviation SEM standard error of the mean sEMG surface electromyography SpO2 oxygen saturation VAB abdominal volume VCW chest wall volume VeeCW end-expiratory chest wall volume VILI ventilator-induced lung injury VRC rib cage volume VT tidal volume Xrs respiratory system reactance ZEEP zero end-expiratory pressure Zin respiratory system input impedance Zrs total respiratory system impedance ΔP driving pressure

Introduction

During more than fifty years of mechanical ventilation, we have learned how to save the lives of patients with respiratory failure and under conditions involving the concurrent failure of other organs. With time, we received proof that we were using a double-edged sword. The severity of complications of mechanical ventilation in terms of morbidity and mortality is com- parable to the diseases for which the mechanical ventilation is indicated, and prevention of these complications is therefore gaining importance. Protective lung ventilation with limitation of tidal volume (VT) and plateau pressure (Pplat) has been widely applied in the treatment of patients with acute lung injury (ALI) and acute respiratory distress syndrome (ARDS), and has been shown to reduce mortality. Positive end-expiratory pressure (PEEP) that overcomes the challenges posed by the reduction of VT has become an integral part of standard of care. However, the optimal level of PEEP and the best method of its titration are still a matter of debate. There are several methods for selecting PEEP, but the search continues for a non- invasive technique that can be used repeatedly at the bedside, without need for deep sedation or paralysis, and new challenges continue to arise, such as measurement of the size of “baby lungs” or assessment of lung recruitability. There is evidence that mechanical ventilation in uninjured lungs may initiate ventilator-induced lung injury (VILI). However, strategies to protect the lungs are not universally used for management of anesthetized patients in the operating room. The available evidence shows that mechanical ventilation with low VT in previously uninjured lungs in the perioperative period can decrease morbidity but not mortality. If it is only the mortality rate that counts, we have to wait for the answers; but until then, we still have to de- cide how to ventilate our patients. Discussions of ventilation techniques include new questions such as the need to individualize low tidal volume settings and to select subgroups for protective ventilation during the perioperative period, and ways of predicting the risk factors. As mechanical ventilation is a supportive therapy, should it not be non- injurious – that is, protective – all the time? Should we not set VT close to the physiological values that have not changed over the last fifty years, not even over the millennia? It seems to be appropriate to start with low VT and

11 increase it if necessary, since it may be too late to reduce VT when there are already complications. The arguments mentioned above, together with the high incidence of postoperative pulmonary complications (PPC) and the high mortality rate of ARDS patients, suggest that attention must be paid to the optimization of ventilation. They also provide an incentive for the use of new techniques and technologies that may contribute to the improvement of respiratory care. The complexity of this issue involves more than just technical develop- ments or the current evidence. There is also knowledge translation, including adherence to the evidence but also the requirement to change our behavior and attitudes, and to change knowledge we have long believed to be correct. This complex process will inevitably be slow, but its progression may be aided by recent and future studies.

12 Background

Mechanical ventilation and ventilator-induced lung injury Since the use of mechanical ventilation began in the early 1960s the range of indications has expanded, both for respiratory failure and for conditions involving failure of other organs, such as head injury, stroke, or liver failure. The goal of mechanical ventilation is to improve oxygenation, ventilation, and lung mechanics while preventing complications. Non-protective ventilation – that is, mechanical ventilation with high tidal volumes (VT), defined 1,2 as VT > 10 ml/kg – may induce VILI. Four mechanisms of VILI have been described. 1) High tidal volume causes overdistention and results in volutrauma, while 2) increasing alveolar pressure leads to barotrauma. 3) Repetitive tidal recruitment with reopening and closing the alveoli throughout each breath causes atelectotrauma, which may induce 4) activation of inflammatory cascade, resulting in biotrauma. The activation of inflammatory cytokines may lead also to dysfunction of the other organs.3,4 Taking into account that mechanical ventilation is a supportive rather than curative therapy, providing time while the disease is treated or is re- solved, the avoidance of complication such as VILI is clearly important.

Protective lung ventilation

A low VT ventilation strategy reduces alveolar overdistention and has been shown to reduce mortality from ARDS.5-7 Another group of strategies, the so-called “open lung strategies”, try to recruit partially collapsed lungs using recruitment maneuvers (RM) and keep the reopened regions open with PEEP. There is increasing evidence that lung recruitment could contribute to VILI, 8-10 due to overdistention of compliant parts of the lungs. Today, low VT and PEEP are considered the standard of care for patients with ALI and ARDS. Low VT here means a volume of approximately 6 ml/kg. However, the findings of Hager and Terragni suggest that there might not be safe Pplat and VT limits in patients with ALI and ARDS,10,11 given the fact that ventilation takes

13 place only in “baby lungs”. It has become obvious that there is no standard approach to determining an optimal PEEP level. Current knowledge on PEEP levels for intensive care patients is based on three studies comparing an aggressive and a conservative PEEP approach (Brower et al., Meade et al., Mer- cat et al.).12-14 All three of these studies showed that a more aggressive strategy improves oxygenation, reduces fraction of inspired oxygen (FIO2) exposure, and improves compliance compared with a conservative strategy. As yet, though, no trial has shown a mortality benefit. Two of these trials have shown results in favor of aggressive PEEP treatment in a subpopulation of patients, but there is still a lack of evidence supporting routine application of high levels of PEEP and ways of determining optimal level. Collapsed alveoli opened by a RM tend to recollapse without adequate PEEP. In other words, RM followed by inadequate PEEP creates unstable alveoli. The resulting cyclic recruitment/derecruitment during mechanical ventilation may contribute to VILI despite improved oxygenation. Too-high PEEP levels can contribute to VILI due to overdistending. Taking into consideration that alveolar injury is often quite heterogene- ous, appropriate PEEP in one region may be suboptimal in another, and ex- cessive in yet another. Therefore, rather than asking whether a patient needs high or low PEEP, we should be looking for optimal PEEP levels which are neither low nor high, but are high enough to keep the maximum number of alveolar structures open throughout the entire respiratory cycle without overdistending them. The problem faced in everyday clinical practice is which PEEP level is optimal for the individual patient, and how it should be titrated. Moreover, the effects of PEEP depend on lung recruitability, which is difficult to investigate bedside. Importantly, Gattinoni et al. observed a correlation between mortality and lung recruitability as demonstrated with computed tomography (CT).9 There are several methods for selecting PEEP: tables, maximal PEEP while avoiding overdistention, gas exchange, stress index, esophageal pressure, pressure-volume curves, and imaging. CT provides an objective tool for assessing lung aeration and consequently for establishing optimal ventilation settings.15 However, CT is expensive and impractical, since it is not available at the bedside, and therefore cannot be used for continuous monitoring. Furthermore, it is associated with high doses of ionizing radiation. In clinical practice, PEEP is usually adjusted according to oxygenation, but this approach may overlook PEEP-induced overdistention as well as intra-tidal recruitment/derecruitment. Also, it is not always matched with lung aeration. One promising approach to setting PEEP level is the assessment of dynamic respiratory mechanics. Dynamic compliance (Cdyn) and respiratory system elastance have both been used as bedside tools for optimization of 16,17 PEEP, and Cdyn has shown correspondence with the findings of CT. It has been suggested that PEEP levels should be set to maximize Cdyn during a

14 decremental PEEP trial. Even though Cdyn has the advantage of being continuously provided by a ventilator, its estimation is strongly affected by non- linearities and within-breath changes in lung mechanics as well as by respiratory muscle activity, requiring either deep sedation or the use of an esophageal balloon in the presence of spontaneous breathing. A possible improvement in assessing dynamic respiratory mechanics and overcoming these limitations is provided by the forced oscillation technique (FOT), which is based on application of a low-amplitude and high frequency pressure forcing signal superimposed on the regular tidal ventilation.

Forced oscillation technique The forced oscillation technique (FOT) is a simple and minimally invasive method for studying the mechanical properties of the respiratory system. The technique stimulates the respiratory system by forcing pressure and simultaneously measures its response to this stimulus in terms of impedance. The forced oscillation is applied at the airway opening, while the measurement of pressure and flow, required for estimation of respiratory system input impedance (Zin), may be performed at this location or more distally. Total respiratory system impedance (Zrs) is based on two parameters: respiratory system resistance (Rrs) and respiratory system reactance (Xrs). Xrs, is related to the respiratory system compliance (Crs) and inertance (Irs), as fol- lows:

�� = 2. �. �. �� − 1/(2. �. �. ��) where f=the oscillatory frequency. Inertance is mainly located in the first gen- eration of airways, which are not affected by recruitment/derecruitment. For this reason, Irs can be considered constant through different conditions, and hence can be neglected when aiming to study relative changes. To quantify lung volume recruitment/derecruitment, a parameter Cx5 (oscillatory compliance derived from Xrs measured at 5 Hz using FOT) can be defined as:

��5 = −1/(2. �. 5. ��)

It has been shown that Xrs measured at an oscillatory frequency of 5 Hz is sensitive and specific to changes in peripheral lung mechanics.18 The ra- tionale behind the method is that the impedance measured using FOT is a measure of the mechanical properties of the airway proximal to the collapsed airway and alveoli, because a change in pressure cannot be transmitted distally past closure and only the proximal airways are oscillated. As airway wall compliance is one order of magnitude greater than lung compliance, Xrs

15 is reduced when a number of alveolar units are prevented from oscillating because of alveolar collapse, Xrs. The strong correlation between Cx5 or reactance measured by FOT and percentage of non-aerated lung tissue suggests that FOT could be used for non-invasive monitoring of lung mechanics and for optimizing ventilator strategy.19 The most important advantage of this approach is that FOT measures respiratory mechanics by applying very low amplitude oscillation to minimize the effects of non-linearities. Moreover, the high frequency used for the forcing signal allows description of the variation of respiratory mechanics within a single breath, allowing measurement at a given lung volume independently from the ventilator pattern.

Experimental setup

Low-amplitude pressure oscillation (∼1.5 cmH2O peak to peak) at 5 Hz was generated by a loudspeaker connected to the inspiratory line of a conventional mechanical ventilator and applied at the inlet of the endotracheal tube (Figure 1). The rear of the loudspeaker was enclosed in a chamber and connected to the inspiratory outlet of the ventilator by a long tube to withstand the positive pressure generated by the ventilator. A digital-to-analogue board mounted on a personal computer generated the signal, which was amplified and played through the loudspeaker. Flow at the airway opening was measured by a pressure transducer connected to a mesh-type heated pneumotachograph. Pressure was measured distally at the tip of the endotracheal tube. All the signals were sampled at 200 Hz by the same A/D-D/A board which controlled the loudspeaker, and were recorded on a personal computer.

Expiratory line Pneumotachograph Mechanical Venlator Inspiratory line

Loudspeaker

Vaoao PPtr

Power ampliﬁer A

A/D-D/AA/D D/A board

Figure 1. Experimental setup. CT – computed tomography; Ptr – pressure; Vao – flow at the airway opening

16 Laser interferometry Another method for the evaluation of oscillatory mechanics is the analysis of how a stimulus (generated in our study by FOT) is propagated through the respiratory system by measuring variations in the displacement of the chest wall surface. This approach allows estimation of the transfer impedance (Ztr). Recently, measurements of chest wall displacement have been success- fully performed by means of an optical sensor based on a self-mixing laser interferometer (LIR), which allows the non-invasive measurement of low- frequency vibrations for the study of oscillatory mechanics of the respiratory system. The introduction of LIR minimizes the main problem in the chest wall estimation; that is, reliable and contactless measurement of its displacement due to both the small amplitude vibrations caused by pressure stimuli and movement due to breathing. Interferometers are used to measure displacement of acquired points of the chest wall to estimate the phase shift between applied pressure at the airway opening and the propagation velocities.

Figure 2. Optical unit. Each black box contains an interferometer.

Experimental setup The chest wall movement was measured in response to a waveform at frequency of 5 Hz. The peak-to-peak amplitude of the stimulus was 2 cmH2O. Interferometry was used to quantify the relative displacement of the measurement points. A total of 20 measurement points were chosen along vertical lines (the midline, the nipple lines, and two parasternal lines) equally distributed from the clavicle to the level of the anterior superior iliac spines. For the experiment, we used a scanning system (Figure 2) consisting of five self-mixing laser interferometers, which were moved by a stepper motor triggered on the respiratory pattern of the patient to cover the chest wall surface as illustrated in Figure 3.

17 AD/DA board computer

interferometers pressure and stepper ﬂow sensor motor expiratory line inspiratory line venlator

loudspeaker

Figure 3. Experimental setup.

Effects of anesthesia on respiratory function and anesthesia-induced atelectasis Almost all anesthetics lower respiratory muscle tone, leading to a fall in functional residual capacity (FRC).20,21 This may reduce airway dimensions and have an effect on chest wall configuration and breathing pattern. The reduced airway dimension, together with mucus stagnation, promotes airway closure, which together with subsequent absorption of trapped gas may lead to alveolar collapse – atelectasis. Preexisting pathology, obesity, postural changes, and type of surgery may enhance these changes and lead to serious consequences during anesthesia or in the postoperative period. Apart from hypoxemia due to a failure to ventilate or intubate a patient, the main causes of gas exchange impairment during general anesthesia that may result in hypoxia are atelectasis formation resulting in shunt, and reduction of FRC. Normally, hypoxemia does not become a clinical problem because it can be very easily compensated by increasing FIO2; on the other hand, several stud- ies22,23 show that a high inspired oxygen concentration is an important factor influencing atelectasis formation during anesthesia. A decrease in inspired oxygen concentration to 80% or 60% during preoxygenation and induction of anesthesia reduces atelectasis during early anesthesia,23 and use of 30% oxygen almost eliminates atelectasis up to 40 minutes after induction.22 The question is, how acceptable is this practice compared with a standard procedure in patient with healthy lungs and no risk factors for difficult airway? Preoxygenation with reduced FIO2 should be instituted only after individual evaluation of risk factors because severe hypoxemia may also occur.24

18 Moreover, 30-40% oxygen, which has been considered standard during anesthesia, has been challenged by studies showing that FIO2 of 0.8 compared with FIO2 of 0.3 reduces the incidence of postoperative nausea and vomiting25 and halves the incidence of surgical wound infection during colo- rectal resection.26 Atelectasis can occur at all ages from infancy27 to old ages.28 It may occur before, during, and after induction of anesthesia,29,30 and is probably due to a combination of different mechanisms such as compression, absorption, loss of surfactant, and other factors such as high FIO2 or obesity. Atelectasis de- velops with both intravenous (i.v.) and inhalational anesthesia and with the patient breathing spontaneously or paralyzed and ventilated mechanically.31 The only anesthetic so far tested that has not produced atelectasis is ketamine,32 although this has also produced the condition when paralysis was used. Atelectasis occurs in 85-90% of patients.28 Up to 20-25% of the lung tissue may collapse, predominantly that located in dependent parts,29 and almost half of the lung can be collapsed after cardiac surgery.33 Atelectasis remains for some time postoperatively depending on surgery and the body mass index of the patient,30 and may persist for days after surgery.3,34

Anesthesia and postoperative pulmonary complications Anesthesia is usually a safe procedure, with very low mortality attributed to the procedure per se. However, it is regularly accompanied by impairment of gas exchange that may result in hypoxia. Observational studies have shown that transient mild desaturation during anesthesia is common, and that more long-lasting and severe hypoxemia may occur with an incidence as high as 20%.35 Around 30% of hypoxemic events occur during emergence and postoperatively.35,36 PPC such as atelectasis, hypoxemia, and pneumonia are relatively frequent and can be associated with increased morbidity, mortality, and economic burdens.37-40 Atelectasis is not the only cause of all postoperative hypoxic events, and there is no direct correlation between atelectasis and pneumonia. Taking into account the number of anesthetic procedures performed worldwide, however, even a small fracture of complications can affect a large number of patients. Despite the lack of conclusive evidence, it seems likely that preventing or actively reversing anesthesia-induced atelectasis could reduce the incidence of PPC. Accordingly, avoidance of high FIO2 during induction and maintenance of anesthesia, as well as intraoperative ventilator settings incorporating restoration of FRC41 and early application of continuous positive airway pressure (CPAP) after major surgery,42-44 has been proposed in patients at high risk of PPC.

19 Lung-protective ventilation strategy during anesthesia and the perioperative period A protective ventilation strategy has been widely applied in treatment of intensive care patients, but is not universally used in the management of anesthetized patients. However, there is increasing evidence that ALI may occur in patients with healthy lungs after onset of mechanical ventilation with a high 45 VT. According to Gajic et al. up to 6.2% of patients without previous lung injury may develop ARDS within 48 hours of initiation of mechanical ventilation with non-protective settings.46 The question thus arises of whether ARDS is an iatrogenic disease caused by mechanical ventilation, or whether mechanical ventilation is responsible for its progression.46-48 The “lung-noxious” process may be aggravated by other risk factors that patients are exposed to during the perioperative period, such as fluid over- load, the inflammatory response to major abdominal surgery, cardiopulmonary bypass, transfusion, and sepsis. A multiple hit theory has been proposed, based on a chain reaction between a predisposing event and various modifying factors; different “hits” can initiate a pulmonary and systemic inflammatory response and may be responsible for the development and progression of ALI/ARDS during the perioperative period.49,50 Once the lung has been primed by an initial insult, injurious mechanical ventilation is an additional hit. Cumulative action of pulmonary insults may explain why mechanical ventilation with non- injurious settings can continue to aggravate lung injury. There is evidence that the pathophysiology of VILI is complex and depends not only on the type of insult but also on the intensity and duration of its exposure. There is also evidence that the same mechanisms of injury in the ARDS lung may apply to mechanically ventilated patients with healthy but partially collapsed lung, which means that atelectasis may contribute to lung injury.50,51 So, theoretically, almost all anesthetized patients are at the risk of lung injury due solely to the presence of atelectasis. The beneficial effect of a lung protective ventilation strategy with low VT and the use of PEEP has been shown in several randomized controlled trials in terms of improved respiratory mechanics, improved oxygenation, shorter duration of mechanical ventilation, decreased incidence of pulmonary dysfunction, and reduced reintubation rate, especially after long procedures.52-56 High VT has been shown to be an independent risk factor for organ failure, multiple organ failure, and prolonged intensive care stay. Recent meta- analyses have shown that protective lung ventilation in previously uninjured lungs in the perioperative period can decrease the development of ARDS, pulmonary infections, and atelectasis but not mortality.57-59 Most of these studies involved patients undergoing cardiac or thoracic surgery, which in itself considered a risk factor for lung injury. Evidence for the use of low VT

20 during low-risk elective surgery is however less secure.60-62 In a recent study, the combination of low VT, RM, and PEEP in patients undergoing abdominal surgery under general anesthesia was associated with improved clinical outcome.63 In contrast, a large multi-center study failed to show any protection from postoperative complications when either low or high PEEP was used.64 However, the latter study did not take into account the large variability of lung mechanics in individual patients. There is thus a need for further studies concerned with the optimization of mechanical ventilation, such as individualization of PEEP level. In the available evidence, zero end- expiratory pressure (ZEEP) is associated with increased hypoxemia, increased rate of iatrogenous-associated infection, and mortality.65-68 In patients without ALI/ARDS, PEEP levels between 5 and 12 cmH2O have been used with low VT ventilation, to avoid atelectasis. Based on the above, many authors suggest the generalization of a protective lung strategy prophylactically to almost all mechanically ventilated patients. Schultz proposed individualization between patients who do not have ARDS but remain at risk for ALI/ARDS, stating that they should be ventilated with VT of 6-8 ml/kg predicted body weight (PBW), while those without risk factors should receive less than 10 ml/kg PBW to prevent lung injury.69 It is known that patients undergoing high-risk surgery and intensive care patients benefit the most from prophylactic protective ventilation.4,46,47,53,55,56,70,71 Woman and obese patients are at higher risk, as they may be particularly exposed to high VT if care is not taken to choose a safe VT. PBW, which is based on height and sex, should be used rather than actual body weight (ABW) for calculation of VT, as the PBW is a better predictor of lung size while the ABW may overestimate the required VT. New information entails the need to respond to issues relevant to clinical practice. When are the healthy lungs at risk of injury? Do healthy lungs need prophylactic protective ventilator settings? Do we need to individualize low VT settings? Although we are dealing here with a population with healthy lungs, at least at the beginning of the process, the complexity of the perioperative period and the multi-hit theory suggest that all ventilated patients may be considered at risk for lung injury. There are two more points that should be taken into consideration. First, early recognition of lung injury in the perioperative period is very difficult, and injury may only be recognized hours or days later.72 Second, it is difficult to predict how many insults are required to cause injury for any particular patient, and it is also a challenge to define which insult is the final one leading to the initiation of lung injury. Therefore efforts should focus on limiting risk factors throughout the whole perioperative period for all patients. One goal of mechanical ventilation is to prevent complications, and it may be argued that ventilator settings should always be protective, including VT very close to the physiological value.

21 There are some challenges to the use of protective ventilation, since low VT can lead to atelectasis and cause hypoxia. Since PEEP can commonly over- come these complications, the combination of low VT and PEEP is widely accepted. PEEP keeps the alveoli open, and setting an optimal level may minimize atelectotrauma, counteract alveolar derecruitment and subsequently hypoxemia and minimize FIO2, which in turn may contribute to the prevention of denitrogenation atelectasis. Application of PEEP is associated with improved oxygenation and morbidity.12,13,73 Multifactorial etiology and high incidence of atelectasis imply the inabil- ity to completely avoid formation of atelectasis. Nevertheless, given the above mentioned early reversal or reduction of anesthesia-induced atelectasis based on an open lung concept during the whole anesthesia and if possible until mobilization, lower FIO2 and particularly a protective ventilation setting with low VT, based on PBW should be considered at least in patients at risk for ARDS. It should perhaps also be considered for all patients during anesthesia, because avoiding ventilator-associated lung injury of the healthy lungs during elective surgery has the same (if not higher) priority than avoiding the same injury in ALI/ARDS patients. Inconclusive results from multiple studies, the high incidence of PPC associated with increased risk of in-hospital mortality,39,74 and the vast number of anesthetic procedures performed worldwide all suggest that attention should be paid to the optimization of mechanical ventilation in anesthetic practice. They also provide an incentive for new techniques, which may contribute, to the improvement of perioperative respiratory care. Early techniques using surface sensors applied to the chest wall suffered from technical limitations such as zero drift that made continuous recording less reliable.75 Another more recent surface sensor technique, electric impedance tomography, represents changes in ventilation rather than volume. A technique that overcomes these limitations is optoelectronic plethysmography (OEP). OEP was developed to simultaneously study volume changes in the thorax and the abdomen, with a temporal resolution and accuracy that was previously unavailable. We used this technique in two studies for continuous assessment of changes in chest wall volume (VCW), and configuration and motion during the perioperative period.

Optoelectronic plethysmography Optoelectronic plethysmography (OEP System®, BTS Bioengineering, Mi- lan, Italy) is a non-invasive method for analysis of chest wall motion. It allows highly accurate measurement of VCW changes of different respiratory compartments in various postures and under various conditions with no need for any individual calibration for the specific subject under analysis. The

22 OEP method cannot, however, measure the absolute lung volume unless the subdivisions of lung volume are known. The volume measurement obtained with OEP, combined with pressure and airflow measurements, can be used to study statics and dynamics of the chest wall and also to estimate possible blood volume shifts. OEP analyzes the movement of 45-52 retro-reflective markers using six television cameras connected to an automatic motion analyzer. Markers are composed of plastic hemispheres of 6 mm diameter covered with thin retro- reflective paper, secured by bio-adhesive, hypoallergenic tape. All markers have to be simultaneously visible to at least two cameras so that their posi- tion and three-dimensional displacement can be reconstructed by stereopho- togrammetry. An image processor measures the positions of each of the markers up to 100 times per second. For volume computation, OEP approx- imates the chest wall surface by a set of triangles connecting the markers (Figure 4). The markers are placed in order to define three chest wall compartments: the upper rib cage, the lower rib cage, and the abdomen. The two rib cage compartments are separated by a horizontal line of markers placed at the level of the xiphoid process. The boundary between the lower rib cage and the abdomen is defined by the markers positioned along the lower costal margin. The total VCW equals the sum of the volumes of the rib cage and the abdomen. A detailed description of this technique has been published else- where.76,77 The method was validated by comparing the lung volume changes obtained by spirometry or pneumotachography with the chest wall total volumes obtained by OEP during different maneuvers.78-83

Motion analysis Model of chest wall

Volume computing

VCW=VRC+VAB

Figure 4. Principles of measurement using opto-electronic plethysmography (OEP). AB – abdomen; RC – rib cage, CW – total chest wall; VCW, VAB, VRC – volumes of the respective areas.

23 Aims

The specific aims of the studies were:

Paper I • To study the effect of propofol anesthesia on chest wall motion during anesthesia induction, spontaneous breathing, and different modes of positive pressure ventilation.

Paper II • To evaluate the effects of three interventions (RM followed by PEEP in combination with moderate FIO2 until extubation) on chest wall volume changes, FRC, and oxygenation, and to compare this approach with a control group in which these interventions were not performed.

Paper III • To evaluate the effects of repeated PEEP optimization based on Xrs, measured by FOT, on oxygenation, lung mechanics, and histologic markers of lung injury, and to compare them with the results obtained by applying the ARDS Network protocol based on oxygenation alone, in a porcine surfactant-depletion lung injury model.

Paper IV • To evaluate the feasibility of using FOT and LIR to measure lung mechanics in healthy anesthetized patients.

24 Materials and methods

The patients, animals, and methods used in the four studies are fully described in the separate study reports (Papers I-IV), to which the reader is referred for details. A brief summary is presented below.

Papers I and II Both studies were reviewed and approved by the Uppsala Human Ethical Review Board. Written informed consent was obtained from all the participating patients.

Patients The populations consisted of 16 (Paper I) and 18 (Paper II) patients of class I-II under the American Society of Anesthesiology (ASA) system, scheduled for elective ear, nose, throat or plastic surgery requiring general anesthesia. Exclusion criteria were age < 18 years, co-existing respiratory disease, antic- ipated difficulties with ventilation or intubation, and significant chest ab- normality.

Anesthesia No premedication was given. Routine monitoring was applied. An arterial line for blood sampling and invasive blood pressure monitoring was placed after induction of anesthesia.

Paper I Total intravenous anesthesia was induced by propofol 2-5 mg/kg (with additional boluses to achieve apnea) and maintained by continuous infusion of propofol 6-9 mg/kg/h in oxygen and air (FIO2 = 0.3 except for 2 minutes before and during anesthesia induction, when FIO2 = 1.0). A size 4 or 5 laryngeal mask airway (LMA) was inserted. After insertion of the LMA and establishment of spontaneous breathing (SB), pressure support ventilation (PSV) was initiated. The pressure support was titrated to achieve an adequate VT (5-10 ml/kg) that resulted in pressure support levels of 4-10 cmH2O. Neuromuscular blockade was achieved using rocuronium 0.6

25 mg/kg, and ventilation was switched to pressure-controlled ventilation (PCV) with the pressure settings at 7-14 cmH2O to obtain similar minute ventilation. The PEEP level was set to zero throughout the study. FRC was measured using the nitrogen washout method.

Paper II An arterial line for blood sampling and invasive blood pressure monitoring was placed after induction of anesthesia. After pre-oxygenation with FIO2 of 1.0 over 2 minutes, a target-controlled infusion was induced, aiming for a target blood concentration of remifentanil 5-8 ng/ml and propofol 3-5 µg/ml. Infusions were reduced to 5 ng/ml of remifentanil and 3 µg/ml of propofol after endotracheal intubation, which was done without use of paralytic agent. For emergence from anesthesia, the infusions of remifentanil and propofol were reduced to 1ng/ml and 1 µg/ml, respectively, to facilitate the return of SB.

Protocol Respiratory movements and thoracic and abdominal volume changes were recorded using OEP. The chest wall was modeled in three compartments: the upper rib cage, the lower rib cage, and the abdomen.

Paper I Volume changes of the chest wall were monitored continuously by OEP throughout a sequence of events comprising pre-oxygenation, induction of anesthesia, SB, PSV and PCV (Figure 5). FRC was measured at three points in the protocol: at pre-induction (quiet breathing), and at post-induction during PSV and PCV.

QB apnea SB PSV PCV

FRC FRC FRC

anesthesia LMA insertion induction

Figure 5. Tracing of the chest wall volume (VCW) in a representative patient for the complete experimental period. LMA – laryngeal mask airway; PCV – pressure- controlled ventilation; QB – quiet breathing; SB – spontaneous breathing.

26 Paper II A pre-oxygenation period of 2 minutes was followed by the induction phase. When patients stopped breathing, no manual ventilation on mask was performed, and they were kept apneic until intubation. Immediately after con- nection of the endotracheal tube to the circuit, FIO2 was reduced to 0.4 while maintaining a fresh gas flow of 10 l/min. Patients were mechanically ventilated with PCV with pressure adequate to reach a VT of 7 ml/kg, a respiratory rate of ten breaths per minute, Ti:Te of 1:2, and PEEP set to zero (although the ventilator had a built-in PEEP of 3 cmH2O). A Draeger Primus ventilator (Drägerwerk AG, Lübeck, Germany) was used initially. Approxi- mately 5 minutes after intubation, the ventilator was switched to an Eng- ström Carestation ventilator (GE Healthcare, USA), which was used for FRC measurements throughout the protocol events. The ventilator parameter settings were identical to those of the Primus ventilator except during FRC measurements, when FIO2 was gradually increased and then decreased according to a built-in protocol.84 OEP data were monitored continuously throughout the different phases of the protocol (Figure 6): 1) awake quiet breathing, 2) induction, 3) onset of ventilation, 4) randomization and intervention (RM), 5) end of surgery and intervention (change of FIO2), 6) spontaneous ventilation during emergence from anesthesia before extubation, and 7) spontaneous breathing of awake patient after extubation. FRC measurements and arterial blood gas sampling were performed at steps 3, 4, and 5 of the protocol. Surface electromyography (sEMG) was performed at steps 1 and 6 of the protocol. The patients were randomly and equally allocated into either the intervention group or the control group. The intervention group received RM (peak -1 pressure 15 cmH2O above 15 cmH2O, I:E ratio 1:2, and frequency 4 min applied for 60 seconds) 5 min after intubation followed by PCV with PEEP of 7 cmH2O and FIO2 of 0.4 during the remainder of anesthesia until the return of SB. At that point, an adjustable pressure limiting (APL) valve was used to maintain CPAP of 7 cmH2O for the spontaneously breathing patient still on FIO2 of 0.4 until extubation. The control group received no RM and no PEEP, and FIO2 was increased to 1.0 5 min before emergence from anesthesia. No CPAP was applied on return of SB with APL valve left fully open.

27 Before Anesth preRM postRM preEME EME postEXTUB 1h later

OEP 0 OEP 1 OEP 2 OEP 3 OEP 4 OEP 5 OEP 6 FRC 1 FRC 2 FRC 3 ABG 4 ABG 1 ABG 2 ABG 3

spontanoeus PCV EMERGENCE spontanoeus breathing breathing ⁄⁄

InterventionIntervention PEEP 7 cm H20 FiO2 0,4

ControlCo to PEEP 0 cm H20 FiO2 0,4 FiOFiO2 1,01,0

⁄⁄ intubaon surgery extubaon

TCI start randomizaon TCI stop preoxygenaon

Figure 6. Protocol flowchart. ABG – arterial blood gas; EME – emergence; FIO2 – fraction of inspired oxygen; FRC – functional residuel capacity; OEP – optoelectronic plethysmography; PCV – pressure-controlled ventilation; SB - spontaneous breathing; TCI – target-controlled infusion

Data analysis In Paper I we calculated the end-expiratory and end-inspiratory volumes of the entire chest wall and its compartments and the contribution to VT of the rib cage and the abdomen, respectively. The phase shift between the upper ribcage and abdominal volume variations was calculated based on the degree of hysteresis of the loop produced when these two volumes were plotted against each other. As a rule, positive angles represent a situation in which rib cage expansion precedes abdominal expansion. In Paper II, minute ventilation, respiratory rate, total and compartmental tidal volumes, and end- expiratory chest wall (VeeCW) variations were recorded during the different phases of the protocol. To estimate the electrical activity of the inspiratory and expiratory rib cage muscles, two disposable Ag/AgCl electrodes connected to a wireless surface electromyographic system, (BTS FREEEMG; BTS, Milan, Italy) were placed in the 8th intercostal right space at the midclavicular line.85

28 Paper III The study protocol was reviewed and approved by the Uppsala Ethics Committee on Animal Experiments.

Animal model and preparation Seventeen pigs (weight 26.6 ± 2.2 kg) of Swedish mixed country breed were included in the study. Anesthesia was induced with tiletamine 6 mg/kg, zo- lazepam 6mg/kg, and xylazine 2.2 mg/kg by intramuscular injection, and maintained with i.v. infusion of phentobarbital 1 mg/ml, pancuronium 0,032 mg/ml and morphine 0.06 mg/ml at the rate of 8 ml/kg/h. After a bolus injection of i.v. fentanyl 10 µg/kg, a tracheotomy was performed and animals were ventilated through a shortened 8 mm inner diameter endotracheal tube in volume-controlled mode (VT 6 ml/kg, PEEP of 5 cmH2O), with respiratory rate titrated to obtain normocapnia. Lung injury was induced by repeated broncho-alveolar lavage with approximately 25 ml/kg warm saline solution per lavage. The endpoint was sustained reductions in the ratio of partial pressure of arterial oxygen (PaO2) to FIO2 (PaO2/FIO2 < 100 mmHg) during a period of 60 minutes. Monitoring according to the standards of our laboratory was established and maintained throughout the experiment.

Protocol The animals were randomized into two groups. Animals in the first group were treated with open lung PEEP (PEEPol) according to Xrs (FOT group). Animals in the second group were treated with PEEP adjusted according to the ARDSNet protocol.6 The duration of the protocol was 12 hours, with every experimental session involving two animals, one from each group. In both groups, RM was performed every 2 hours by increasing PEEP to 20 cmH2O for 2 minutes, preceded by tracheal suctioning for 5 seconds, to sim- ulate a clinical situation. In the FOT group, PEEPol according to Xrs was identified during a decremental PEEP trial, performed immediately after a RM by a stepwise reduction of PEEP from 20 cmH2O in steps of 2 cmH2O until Xrs reached its maximum and started to decrease (Figure 7). PEEPol was defined as the PEEP at the step preceding the first reduction of Xrs. Immediately after obtaining PEEPol, PEEP was increased to 20 cmH2O for 1 minute and then brought back to PEEPol and maintained for the next 2 hours. Both groups received the same number of RMs. Respiratory rate and FIO2 were adjusted according to the ARDSNet protocol in both groups. After the animals were sacrificed, thoracotomy was performed, and sections from the left lingual and left lower lobe (two sections from each lobe) were prepared for the histopathological analysis.

Figure 7. Positive end-expiratory pressure (PEEP) optimization procedure, according to optimal respiratory system reactance (Xrs). The upper panel shows tracheal pressure and the lower panel shows Xrs measured at end expiration.

Data analysis

The groups were compared in terms of ventilator parameters (PEEP, Cdyn, Pplat), driving pressure (ΔP), gas exchange, and hemodynamics. Estimation of Zrs was obtained from the flow and pressure signals. Total respiratory system impedance was expressed as real part (Rrs) and imaginary part (Xrs). Four fields for each animal were evaluated randomly for the histopathological analysis, two from the lingual and two from the left lower lobe. A grading scale (0-4) for four different histopathological markers of lung injury was used: presence of alveolar edema, hyaline membranes, and inflammatory cells in alveoli and in septa, respectively, (modified from Akinci et al.).86 The evaluation scores from these markers were averaged to obtain a cumulative histopathology score for each animal.

Paper IV The study was approved by the Uppsala Human Ethical Review Board. Written informed consent was obtained from all the participating patients.

30 Patients The study populations consisted of 15 ASA class I-II patients scheduled for elective ear, nose, throat or plastic surgery requiring general anesthesia. Ex- clusion criteria were identical to those of Paper I and II.

Anesthesia The patients were premedicated with midazolam and paracetamol. Routine monitoring was applied, and an arterial line was placed after induction of anesthesia. Total intravenous anesthesia was induced by propofol 2-3 mg/kg and a bolus of remifentanil 0.3 µg/kg to facilitate the insertion of LMA, while maintaining SB during induction. Anesthesia was maintained by a continuous infusion of propofol 6-7 mg/kg/h. After measurements at this step were completed, infusion of remifentanil 0.02 µg/kg/h was started and endotracheal intubation was facilitated by i.v. administration of rocuronium 0.5 mg/kg. Patients were mechanically ventilated with PCV (Draeger Pri- mus, Drägerwerk AG, Lübeck, Germany) with pressure titrated to reach VT of 8 ml/kg, respiratory rate of 10 breaths per minute, I:E ration 1:2, ZEEP, and FIO2 of 0.4.

Protocol The baseline pulmonary function was assessed by spirometry prior to anesthesia. Subsequently, the patients were studied throughout the different phases of the protocol: 1) awake quiet breathing subject, 2) spontaneous ventilation through LMA after induction, 3) PCV with ZEEP after intubation, 4) PCV with PEEP of 5 cmH2O, 5) RM (peak pressure of 30 cmH2O and PEEP of 15 cmH2O), and 6) PCV with PEEP of 5 cmH2O after RM.

Data analysis For each protocol step the following parameters were measured and recorded: input impedance (Zin) at 5 Hz measured by FOT, displacement of the chest wall assessed by LIR at a frequency of 5 Hz, and arterial blood gas. Dynamic compliance and resistance (Cdyn and Rdyn) were calculated for each steps of the protocol. At each condition, a color map of pressure wave propagation velocities was estimated. From each point at the chest wall, the velocity was extracted and the phase delay between the pressure at the airway opening and each point was estimated as phase of transfer function between pressure and velocities computed by spectral methods.

31 Statistical analysis (Papers I-IV) Paper I One-way analysis of variance (ANOVA) was applied to study the difference in end-expiratory and end-inspiratory volumes, contribution to VT, and phase shift in difference chest wall compartments. Data are presented as mean (standard error of the mean).

Paper II Anthropometric and clinical data of both groups were compared using ANOVA with group as independent factor. In order to compare hemodynamics, the breathing pattern, and VCW variations between the two groups, a two-way ANOVA on repeated measures was performed with the different phases of the protocol and group as independent factor. The difference in VeeCW between the first and the last breath during emergence phase were evaluated using Student’s t-test. Results were considered significant if p<0.05. Data are presented as mean (standard deviation, SD).

Paper III Normality was tested using the Kolmogorov-Smirnov test. Significance of differences between baseline parameters in the groups was tested using an unpaired t-test when the normality test passed, and using the Mann-Whitney test when the normality test failed. Significance of differences between the two groups was tested by two-way ANOVA for repeated measurements using group and protocol steps as factors. Multiple comparisons after ANO- VA were performed using the Holm-Sidak test. Significance of differences between the histopathological scores was tested using the Mann-Whitney test. Results were considered significant if p<0.05.

Paper IV ANOVA with Bonferroni’s multiple comparison test was applied to study the significance of differences in Rrs, Xrs, Cdyn, Rdyn and the values of arterial blood gas and mechanical ventilation. Results were considered significant if p<0.05. Data are expressed as mean (SD).

32 Results

Paper I

Total VCW decreased by 0.41 ± 0.08 L after the induction, but was then partially restored towards baseline levels, remaining at 0.24 ± 0.1 and 0.22 ± 0.1 L below the awake level during PSV and PCV, respectively. The decrease in end-expiratory VCW after induction was almost equally shared between the rib cage and the abdomen. SB after apnea resulted in partial restoration of end-expiratory VCW. End-expiratory VCW was not affected during PSV and PCV (Figure 8).

0.8 0.6 0.4 * 0.2 * end-inspiration * (liters) 0.0

CW -0.2 V * end-expiration -0.4 -0.6 QB apnea SB PSV PCV

0.4 0.4

0.2 0.2 * * * (liters) 0.0 (liters) 0.0 AB RC V V -0.2 * * -0.2

QB apnea SB PSV PCV QB apnea SB PSV PCV

Figure 8. Mean ± standard error of the mean (SEM) values of the end-expiratory (closed circles) and end-inspiratory (open circles) volumes of the chest wall (VCW, top panel), the rib cage (VRC, left lower panel), and the abdomen (VAB, right lower panel) in different conditions of the protocol. ∗ p<0.05 relative to quiet breathing (QB). PCV – pressure-controlled ventilation; PSV – pressure support ventilation; SB – spontaneous breathing.

Figure 9 shows that the abdominal compartment dominated chest wall motion during quiet breathing, but the relative contribution of the upper rib cage increased significantly in transition from SB to PSV. During quiet breathing, analysis showed negative phase shift angles (-17.4 ± 4.9°) between the upper

33 rib cage and the abdomen, suggesting that inspiration was initiated by the diaphragm (Figure 10). The phase shift angle became positive during SB (15.3 ± 6.9°) and PSV (11.3 ± 3.4°), but was lower during PCV (2.3 ± 0.8°), indicating that the intercostal muscles were leading under anesthesia during both SB and PSV (Figure 10).

80 70 60 abdomen 50

40 upper rib cage 30 ** ** 20 lower 10 rib cage

Vol. variation (%tidal volume) 0 QB SB PSV PCV

Figure 9. Mean ± standard error of the mean (SEM) of the volume changes of the different chest wall compartments during different conditions of the protocol, expressed as percentage of the tidal volume (VT). ∗ p<0.05 relative to quiet breathing (QB). PCV– pressure-controlled ventilation; PSV – pressure support ventilation; SB – spontaneous breathing.

30 Vab * 20 Vrcp * Vrcp 10 Time Vab (degrees) 0

-10 Vrcp phase shift phase

-20 Vrcp

Vab -30 Time Vab QB SB PSV PCV

Figure 10. Mean ± standard error of the mean (SEM) of the phase shift angle between upper rib cage (VRCP) and abdominal (VAB) volume variations during different conditions of the protocol. Positive values represent a condition when the rib cage leads the abdomen and a clockwise VAB-VRCP loop is observed. Negative values represent a condition when the abdomen leads the rib cage and a counter-clockwise VAB-VRCP loop is observed. ∗ p<0.05 relative to quiet breathing (QB).

34 Figure 11 shows detailed OEP tracing from different compartments. The upper rig cage initiates breathing, while breathing motion is evident in the abdomen almost 60 seconds later. Full coordination between compartments, resulting in larger VT, is seen still later.

a PERIOD 1 PERIOD 2 PERIOD 3 8.40 Upper rib cage volume (L)

8.35

8.30

8.25

3.90 Lower rib cage volume (L) 3.85

3.80

7.85 Abdominal volume (L) 7.80

7.75

7.70

20.15 20.10 20.05 TOTAL chest wall volume (L) 20.00 19.95 19.90 19.85 19.80 19.75 03060901201501802102 40 time (sec)

b PERIOD 1 PERIOD 2 PERIOD 3

5101520 80 85 90 160 165 170

time (sec)

Figure 11. Volume variation of the upper rib cage (blue), lower rib cage (green), abdomen (red) and total chest wall (black) during the transition from spontaneous breathing in a representative patient.

35 Paper II Two of the 18 patients enrolled were excluded due to missing OEP data, and two patients from the intervention group were analyzed separately due to a violation of the study protocol (specifically, CPAP was omitted during the emergence phase). After the induction of anesthesia, VeeCW decreased progressively with a similar fall in both groups during post-induction apnea (Figure 12). The onset of mechanical ventilation did not introduce any differences in VeeCW between the two groups. After RM and application of PEEP for the remaining phases of the protocol, VeeCW was consistently higher in the intervention group compared to the control group (p<0.001). During the entire protocol, the latter group was characterized by decreased VeeCW compared to awake quiet breathing, without returning to baseline after extubation. In contrast, in the intervention group VeeCW increased significantly after extubation (p<0.001). The effect of RM followed by PEEP disappeared in the two patients (the light gray symbols in Figure 12) in whom CPAP was not applied at the beginning of the emergence phase, and in this subgroup VeeCW followed a pattern similar to the control group thereafter.

Figure 12. Average values ± standard deviation of end-expiratory chest wall volume variations (VeeCW) during the different phases of the protocol. The grey area represents the emergence phase, including the first and last breaths of the emergence phase as well as spontaneous breathing before extubation. Closed symbols represent the intervention group (RM+), open symbols represent the control group, and grey symbols represent the two excluded RM+ patients in whom CPAP was omitted during the emergence. ∗∗∗: p<0.001 vs. control; °°,°°°: p<0.01, 0.001 vs. BEFORE ANESTH.

36 The grey areas in Figures 12-15 represent the emergence phase, which started when propofol and remifentanil infusions were stopped, mechanical ventilation was subsequently switched off, and the patients gradually resumed their spontaneous breathing. During this period, VeeCW tended to decrease with time in both groups although the levels in the intervention group were still above those in the control group (p<0.001). More specifically, as indicated in Figure 13, end-expiratory rib cage and abdominal volumes significantly decreased between the first and last breaths of the emergence phase in all but one patient, regardless of group affiliation and despite the presence of 7 cmH2O of PEEP in the intervention group.

Figure 13. End-expiratory chest wall (left panel), rib cage (middle panel), and abdominal (right panel) volume variations between the first and last breaths of the emergence phase before extubation. Closed symbols represent the intervention group, and open symbols represent the control group. ∗∗, ∗∗∗: p<0.01, 0.001 vs. first breath.

Expiratory time, expressed as a percentage of the total respiratory cycle time, significantly increased during the emergence phase in both groups (p<0.01) and returned to previous waking levels after the extubation (Figure 14). Representative tracing of variations of VeeCW and sEMG signal during awake quiet breathing and emergence are shown in Figure 15. In the subset of patients in whom sEMG was measured, the expiration/inspiration ratio for the root mean square of the sEMG signal (RMS,exp/RMS,insp) ration was 0.95 ± 0.10 and 1.17 ± 0.12 during awake breathing and emergence respectively, indicating an enhanced expiratory activity during the emergence from anesthesia (p=0.008).

Figure 14. Average values ± standard deviation of the expiratory time (expressed as percentage of total respiratory cycle time) during the different phases of the protocol in which the patients were breathing spontaneously: quiet breathing (BEFORE ANESTH), emergence phase (EMERGENCE, grey area), immediately after extubation (PostExt), and one hour after extubation (1h PostExt). Closed symbols represent the intervention group, and open symbols represent the control group. ∗∗: p<0.01 vs. BEFORE ANESTH, PostExt, and 1h PostExt.

Figure 15. Top panels: chest wall volume variations (VCW) and the surface electromyography signal (sEMG) at the 8th intercostal space in a representative patient during quiet breathing followed by a slow vital capacity breath pre-induction (BE- FORE ANESTH) and spontaneous quiet breathing during the emergence phase (EMERGENCE, grey area). Bottom panels: VCW and sEMG of the four highlighted breaths (white area) during the emergence phase. Active expiration occurs during emergence, as revealed by the increased sEMG signal during expiration in the highlighted breaths.

38 Paper III

Figure 16 presents the values of maximal Xrs and the optimal PEEP identified in all animals at the different optimization steps. The increase in Xrs clearly shows that there was an average improvement in the oscillatory mechanics with time, and this led to a progressively lower PEEP applied to the FOT group. The parameters measured every hour during ventilation trial were averaged for all animals. The values of PEEP, Cdyn, Pplat and ΔP are reported in Figure 17, and the values related to gas exchange and hemodynamics are reported in Figure 18.

Figure 16. Time course of positive end-expiratory pressure (PEEP) and respiratory system reactance (Xrs) in the FOT group. Mean ± standard deviation of maximum Xrs values (closed symbols) and optimized PEEP (open symbols) assessed during optimization procedudre.

At the beginning of the trial, optimization based on Xrs resulted in a significantly higher PEEP compared with that set in the ARDSNet group. These settings led to a significantly lower ΔP, a better oxygenation, and lower mean arterial pressure and mean pulmonary arterial pressure in the FOT group. Over the course of the 12-hour experiment, PEEP decreased in both groups from 10.4 ± 1.7 to 8.9 ± 1.8 cmH2O in the FOT group, and from 7.4 ± 2.1 to 5.0 ± 0 cmH2O in the ARDSNet group. These higher values of PEEP in the FOT group were associated with improved respiratory mechanics, as indicated by the significantly lower ΔP (decreasing from 10.1 ± 2.05 to 9.88 ± 1.78 vs. decreasing from 16.9 ± 5.2 to 13.4 ± 4.4 cmH2O in the ARDSNet group) and the higher Cdyn for most of the course of the experiment. There was a trend for lower Pplat in the FOT group, but the differences

39 between the groups were not significant. At the end of the experiment, only changes in oxygenation and PEEP were significantly different.

Figure 17. Ventilatory and respiratory mechanics parameters over time. Positive end-expiratory pressure (PEEP), plateau pressure (Pplat), driving pressure (ΔP), and dynamic compliance (Cdyn) for the FOT group (closed symbols) and the ARDSNet group (open symbols). Data are presented as mean ± standard deviation. ∗, p<0.01; +, p<0.05

After 12 hours of ventilation, histopathological analysis showed a significantly lower score for lung injury in the FOT group compared with the ARDSNet group (Table 1). This is illustrated in Figure 19, with representative sections from both groups.

Figure 18. Blood gases and hemodynamic parameters over time. Partial pressure of arterial oxygen (PaO2)/fraction of inspired oxygen (FIO2), partial pressure of arterial carbon dioxide (PaCO2), mean arterial pressure (MAP), and mean pulmonary arterial pressure (MPAP) for FOT group (closed symbols) and ARDSNet group (open symbols). Data are presented as mean ± standard deviation. ∗, p<0.01; +, p<0.05

Table 1. Lung injury scores for FOT and ARDSNet groups. Data are reported as mean ± SD. ARDSNet FOT p Alveolar edema 0.88 ± 0.92 0.06 ±0.18 0.03 Hyaline membrane 0.94 ± 0.73 0.69 ±0.70 0.50 Alveolar infl. cells 1.31 ± 0.80 0.81 ± 0.46 0.15 Septal infl. cells 2.69 ± 0.80 2.25 ± 0.65 0.25 mean ± SD 1.45 ± 0.47 0.95 ± 0.37 0.03

Figure 19. Histopathology images of representative lung samples from FOT group (left) and ARDSNet group (right). There is more alveolar edema and inflammatory cells in both septa and alveoli in the animal ventilated with PEEP suggested by ARDSNet.

Paper IV The real and imaginary parts of input impedance were estimated for each patient, frequency, and anesthesia stage, respectively, and then were averaged for all the patients. Figure 20 shows the input impedance parameters reactance (Xrs) and resistance (Rrs) at a frequency 5 Hz. Rrs decreased slightly after intubation and even further (3.6 ± 2.9 to 1.3 ± 1.6 cmH2O.s/l; p<0.001) when mechanical ventilation with PEEP of 5 cmH2O was initiated. In contrast, Xrs initially decreased slightly, but subsequently increased from -4.5 ± 3.4 to -2.1 ± 2.0 cmH2O.s/l (p=0.002). Changes in both parameters were negligible at the other steps of the protocol. We observed a positive trend in the changes of Cdyn for each step of the protocol, but significantly higher Cdyn was observed only at the last step with PEEP of 5 cmH2O after RM. Interferometers were used to measure the displacement of each point of the chest wall to estimate the phase delay between applied pressure and the velocities of propagation. From each pressure and velocity, the phase delay was estimated and plotted to produce a color map as illustrated in the colored upper part of Figure 21. The first two rows correspond to the upper and lower rib cage respectively, while the last two measure displacement of the upper abdomen and around the umbilicus line, respectively. Figure 21 shows maps of the phase shift at 5 Hz. At every protocol step, it is evident that the chest wall behaves as a multi-compartmental region; the separation of rib cage and abdomen is the most striking feature. It is possible to identify the region of the rib cage, where the pressure stimulus moves fast, and the abdomen, where the pressure wave is slowed down. The same trends can be traced by plotting the mean of each row for each condition, as illustrated in the lower part of Figure 21. The lower rib cage

42 and upper abdomen show similar trends while the lower abdomen exhibits a different trend. The phase delay during mechanical ventilation shows a min- imum at ZEEP for the lower rib cage and upper abdomen. The values at PEEP of 5 cmH2O increase and remain largely unchanged after RM. Both the color maps and the phase delay values indicate the lower abdomen as the region with different trend.

Figure 20. Respiratory system resistance (Rrs; the upper panel) and reactance (Xrs; the lower panel) analyzed for each subsequent protocol step (including PEEP of 5 cmH2O both before and after RM). Data are presented as mean (standard deviation) values.

2 phase delay(rad) 1 LRC 0 UABD LABD -1 QB LMA ZEEP PEEP5 PEEP5RM

Figure 21. Phase delay at 5 Hz during quiet breathing (QB), anesthesiawith spontaneous ventilation through a laryngeal mask (LMA), pressure-controlled ventilation with PEEP of 0 cmH2O (ZEEP), 5 cmH2O (PEEP5), and 5 cmH2O after RM (PEEP5RM). Above: colored maps of compartmentalized chest wall displacement at the different phases of the protocol. Below: average phase delay values (mean ± standard deviation) for the lower rib cage (LRC), upper abdomen (UAB) and lower abdomen (LAB)

44 Discussion

Main findings The most important findings from the studies included in this thesis are:

Propofol anesthesia decreases end-expiratory chest wall volume, with a more pronounced effect on the diaphragm than on the rib cage muscles. The rib cage muscles initiate post-apneic breathing after induction of anesthesia.

The restoration of breathing during emergence from propofol anesthesia is associated with prolonged expiratory time, activation of the expiratory muscles, and decreased end-expiratory chest wall volume. The combination of a RM, PEEP, and reduced FIO2 may preserve lung volume during and after anesthesia.

In a lavage model of lung injury, a PEEP optimization strategy based on maximizing oscillatory reactance, measured by FOT, resulted in improved lung mechanics, increased oxygenation, and reduced histopathologic evidence of VILI.

Forced oscillation technique and laser interferometry were possible to apply simultaneously for respiratory monitoring of the anesthetized patient.

Papers I and II To our knowledge, the study presented in Paper I is the first to address changes in VCW, configuration, and motion in a full sequence consisting of awake quiet breathing, induction of anesthesia, spontaneous breathing during anesthesia, and different modes of mechanical ventilation; and Paper II presents the first study in which VCW, breathing pattern, and respiratory muscle activation were investigated specifically during the emergence period.

45 The role of rib cage muscles during spontaneous breathing and anesthesia The main novel findings presented in Paper I were an increased relative contribution of the rib cage to ventilation after induction of anesthesia, and the fact that the rib cage initiates post-apneic ventilation; the intercostal respiratory muscles were leading each breath during spontaneous breathing under anesthesia. As shown previously,80,87 the diaphragm was dominant during quiet breathing, but when breathing returned after post-induction apnea, rib cage expansion preceded abdominal expansion. This pattern con- tinued during PSV. If this could be corroborated in further studies, it might have implications for new modes of triggering ventilator support of spontaneous ventilation. In conjunction with the above findings, the rib cage muscles not only initiate each breath in the anesthetized subject during spontaneous breathing, they are also the first to “wake up” after post-induction apnea, while the abdominal compartment begins to move almost 30 seconds later. This is not in agreement with earlier findings that (volatile) anesthetics have more pronounced effects on the rib cage muscles than on the diaphragm.88,89 The results confirm findings from previous studies of rib cage configuration changes and diaphragm displacement during anesthesia.75,90 Further- more, Strandberg et al. observed post-induction end-expiratory volume reduction of 500 ml,31 subsequently restored with return of SB, in accordance with the reversible reduction of over 400 ml found in our study (Figure 8). Interestingly, neither PSV nor PCV with paralysis increased end-expiratory volume further. In the present study, chest wall motion changed on the induction of anesthesia. The rib cage contributed relatively more to the tidal volume, indeed as much the abdominal compartment, during PSV and PCV (Figure 9). This is in contrast to the findings of Tusiewicz et al.88 who reported that halothane anesthesia decreased the relative contribution of the rib cage to VT. These conflicting results may be due to the different anesthetic agent used, corroborating the findings of Brown.75 Two studies have demonstrated the inhibitory effect of propofol on diaphragmatic contractility in patients during anesthesia.91,92 In view of this fact, a study comparing the effects of different i.v. and inhalational agents using OEP would be useful.

Active expiration during emergence from anesthesia The main new finding of Paper II was that the restoration of breathing during emergence from anesthesia was associated with prolonged expiratory time, active expiration, and reduction in VeeCW. We assume that the prolonged expiration was a consequence of the active contraction of the expiratory muscles and speculate that the active expiration was triggered by resistance to deflation of the lungs due to the endotracheal tube. This is sup-

46 ported by the shortened expiratory time after extubation. The results provide new information by quantifying reduction of VeeCW during the relatively short period after surgery when anesthesia and ventilation are discontinued and spontaneous breathing returns. The VeeCW reduction during the emergence phase was consistent regardless of whether RM and PEEP were applied, although PEEP kept rib cage and abdominal volumes elevated compared to the controls. Active contraction of expiratory muscles has previously been demonstrated to lower VeeCW in order to recruit expiratory reserve volume during exercise in healthy humans.93 Interestingly, in the latter study the volume reduction was confined to the abdomen during exercise, while in the present study it was mainly observed in the rib cage compartment. The active expiration might have been due to the expiration reflex, which is characterized by prompt expiratory effort without preceding inspiration, in contrast to the cough reflex, which is characterized by different neuronal pathways and subsequently activation of different muscle groups.94-96 Stud- ies by Nishino et al. have shown that general anesthesia blocks the cough reflex more effectively than the expiration reflex.97,98 Strong expiratory efforts are in conformity with the higher RMS,exp/RMS,insp ratio, the prolonged expiratory time, and the reduction in VeeCW observed during emergence.

The open lung concept

Furthermore, RM in combination with PEEP and FIO2 of 0.4 until tracheal extubation resulted in increased postoperative VeeCW compared with pre- induction conditions, while VeeCW decreased in the patients ventilated without RM and PEEP, and in whom FIO2 was increased to 1.0 during the emergence phase. In the intervention group, the volume reduction during emergence was restored after extubation, in contrast to the control group. Interest- ingly, in the two patients in the intervention group in whom CPAP was omitted during the emergence phase, VeeCW fell to levels approaching those of the control group. This suggests that PEEP is important to prevent atelectasis formation, in particularly if high FIO2 is used during the emergence. It may thus be beneficial to maintain PEEP/CPAP during the emergence phase.

Paper III In Paper III, we followed lung mechanics and ventilation parameters throughout the course of 12 hours, which more closely resembles a clinical situation with time enough for the more subtle mechanisms of VILI to have effect.

47 Although CT is still the gold standard to assess lung volume recruitment, there is increasing evidence that lung mechanics are a better surrogate than gas exchange variations for the assessment of lung recruitment at the bed- 99 16,17,100,101 side. Several studies suggest that the use of Cdyn might guide in the identification of the optimal PEEP. A recent study of ALI/ARDS patients used a combination of oxygenation data and lung volume changes obtained by electrical impedance tomography.102 The authors reported that volume-dependent compliance seemed to be superior to Cdyn over the whole breath for monitoring lung recruitment and defining optimal PEEP. Howev- er, this method is labor intensive and expensive. In addition, the monitoring of esophageal pressure in order to maintain positive trans-pulmonary pressure has recently been suggested for PEEP optimization.103 However, the necessity of an appropriate positioning of the esophageal balloon and the intrinsic difficulties of such a measurement imply problems whit the imple- mentation of this technique in clinical practice. Conversely, FOT allows the peripheral lung mechanics to be continuously monitored via the ventilator circuit, and so this could be a preferable technique. Bellardine et al. applied FOT in an animal model of ARDS to study changes in lung mechanics at different PEEP levels.104 The authors found that optimal PEEP identified by CT scans minimizes mechanical heterogeneity, defined as the frequency dependence of Rrs and elastance. However, this approach requires the assessment of mechanical impedance on a frequency range of 0.2 to 8 Hz, and hence, is not suitable for patients with spontaneous breathing. In two previous studies, we have shown that single-frequency FOT of 5 Hz can be used to accurately evaluate lung volume de-recruitment, overcoming several limitation of Cdyn, such as the effect of non-linearities in the respiratory system and the need for deep sedation or paralysis of the patients.19,105 The optimization of PEEP on the basis of Xrs changes was clearly advan- tageous compared with PEEP settings according to the ARDSNet protocol. However, in the FOT group in which Xrs was continuously monitored, we occasionally observed decreases in Xrs during the two-hour intervals between the scheduled RMs, and no adjustments were made. These results suggest that by monitoring Xrs it is possible to continuously assess the development of lung collapse and to evaluate the efficacy of RMs, allowing bedside char- acterization of lung recruitability.

48 Paper IV In Paper IV, we showed the possibility to apply both FOT and LIR simultaneously in various conditions ranging from awake quiet breathing to general anesthesia with controlled mechanical ventilation. Reactance at 5 Hz (Xrs) has been demonstrated to function as an index of the amount of open and ventilated lung. Changes in PEEP levels caused Xrs to increase, evidencing the presence of lung collapse, but RM in this trial caused no variation in Xrs, suggesting that the degree of initial lung collapse was small and no further recruitment could be obtained. This is not surpris- ing since the patients were healthy and the induction protocol minimized lung collapse, including spontaneous breathing and reduced FIO2. The second method for evaluation of oscillatory mechanics was to analyze how the stimulus was propagated through the respiratory system by measuring the variations in the displacement of the chest wall surface. This approach allows the estimation of the transfer impedance (Ztr). The present paper describes the first use of LIR in anesthetized patients, although previ- 80,106,107 ous studies have attempted to quantify resistance, elastance, and Ztr. The introduction of LIR minimizes the main problem in estimating the chest wall mechanics; that is, reliable and contactless measurement of its displacement due to both the small amplitude vibrations and movement due to breathing. In addition, while measurement of Zin by FOT could be described as the response of a single compartment to small pressure stimuli generated externally, LIR allows the assessment of Ztr in several predefined areas of the chest wall and may therefore be considered as more adequate for detecting heterogeneity and regional changes. By comparing the results along the same row, the data may be considered highly reproducible, since the five interferometers are completely independent.

49 Conclusions

• Propofol anesthesia decreases end-expiratory chest wall volume. It has a more pronounced effect on the diaphragm than on the rib cage muscles, which demonstrate an increased relative contribution to ventilation after induction of anesthesia by multifaceted effects.

• The decrease in end-expiratory chest wall volume during emergence from anesthesia, associated with activation of the expiratory muscles, suggests that active expiration may contribute to FRC reduction during emergence. However, the combination of a RM, PEEP, and reduced FIO2 may preserve lung volume during and after anesthesia.

• Optoelectronic plethysmography is a powerful tool for high temporal and spatial resolution studies of chest wall kinematics in anesthetized patient.

• A PEEP optimization strategy using forced oscillation technique may reduce ventilator-induced lung injury.

• Forced oscillation technique and laser interferometry can be applied simultaneously for respiratory monitoring of the anesthetized patient.

50 Limitations

The studies described in this thesis only included patients with ASA classi- fycations I and II, and so the results may not be extrapolated to, for example, obese patients and patients with respiratory disease, in whom the potential negative effects of anesthesia and controlled mechanical ventilation on respiratory functions could be more pronounced. Furthermore, the conclusions drawn from these studies apply only to anesthesia with propofol or a combination of propofol and remifentanil, as we did not investigate other anesthetic agents which could influence the respiratory muscles in a different way.108 Most of the earlier studies on the effects of anesthesia on the mechanics of breathing studied other agents such as halothane, ketamine, and thiopen- thone using older techniques or CT.31,88,109 Thus, direct comparison between the results of these earlier studies and Paper I must be carried out with cau- tion. It must also be recognized that OEP measures, albeit very precisely, variations in chest wall volume, which is not the same as tidal ventilation per se. Changes in chest wall configuration could be caused by displacement of the diaphragm, the development of atelectasis, the effects of anesthesia on the rib cage muscles, or blood volume shift110-112 from the thorax when intra- thoracic pressure is increased due to pressure ventilation. The individual effects of RM, PEEP, or FIO2 on VeeCW (Paper II) were not elucidated, and further studies are needed to address this issue. The saline lavage model of lung injury used in Paper III causes surfactant depletion and atelectasis that is easily recruitable, in contrast to the hetero- geneous inflammatory changes that characterize ARDS. It could also account for the clinical improvement through the course of the experiment. An advantage of this was that the final damage seen in histopathologic sections could most likely be attributed to mechanical ventilation, the focus of the study, rather the initial lavage injury. The changes in Xrs include changes in both lung and chest wall mechanics, as the esophageal pressure was not measured. Qualitative and semi-quantitative multi-parameter analysis of histopathologic changes was used instead of true quantitative analysis.

51 Future perspectives

The conclusions drawn from the OEP studies apply to intravenous anesthesia with propofol or a combination of propofol and remifentanil. We did not study other anesthetic agents, which could influence the respiratory system via different mechanisms. Consequently, a study comparing the effects of propofol, ketamine and inhalational agent using OEP, which overcomes the limitations of older techniques, would be useful.

Various studies have yielded conflicting results regarding blood volume shifts caused by anesthesia and paralysis.109,113,114 The present OEP studies could not address this issue, which merits future studies investigating blood volume redistribution79,115 during anesthesia, mechanical ventilation, and RM.

FOT can be easily integrated in commercial mechanical ventilators and could provide continuous monitoring of the mechanical properties of the respiratory system at the bedside. The results of the FOT experimental study suggest that by monitoring Xrs it is possible to continuously assess the development of lung collapse; this could be used to create Xrs-based triggers for RM. Further studies should aim to validate FOT and LIR for detecting derecruitment in anesthetized patients, and to determine if LIR also has a high sensitivity in pathological condition.

52 Acknowledgements

I would like to express my sincere gratitude to everyone who has contributed directly or indirectly to this thesis, and in particular to:

Peter Frykholm, my main supervisor, for introducing me to scientific work, for always taking the time to read and discuss our work, for your insightful comments and enormous assistance in creating the draft manuscript, for your enthusiasm, for a rare understanding of what is important in both work and life, and for your modesty and humbleness. Thank you very much.

Anders Larsson, my co-supervisor, for the many constructive suggestions and comments that have improved my work so incredibly much, for your support and pragmatism that saved me when times were hard and obstacles almost insurmountable, and at last but not least, for providing time for research.

Göran Hedenstierna, for enthusiastic support and fruitful discussions, and for all the swift, concise reviews and insightful comments that have improved my work unbelievably much.

Sten Rubertsson, for providing time for research, and for development of my teaching skills when keeping me busy with educating medical students.

Torsten Gordh, for recruiting me, for introducing me to Göran Hedenstierna, and for your kindly encouragement while asking me the same question almost every time when we met: “When are you going to start your research?” When I had no more excuses, I started.

Lars Wiklund, for creating an inspiring research environment at the Depart- ment of Anesthesiology and Intensive Care.

Torsten Gordh, Torbjörn Karlsson, Kristina Hersio and Fredrik Lennmyr, former and present heads of the Department of Anesthesiology and Intensive Care and the Department of Thoracic Surgery and Anesthesia, for creating conditions for research and allowing me the necessary time off from clinical work.

53 Agneta Roneus and Karin Fagerbrink, for your permanent availability and invaluable help at the animal laboratory.

Monica Segelsjö, for your excellent work and assistance in CT scanning late into the evening.

Katja Andersson, for your constantly kind and great assistance in many situ- ations.

All my co-authors and friends at Dipartimento di Bioingegneria, Politecnico di Milano, for the enormous work behind the technical part of these studies, for your patience when trying to induct a physician into the secrets of phys- ics and mechanical modeling of the respiratory system, and for your infinite enthusiasm despite experiment trials lasting nearly the whole day.

All my colleagues at the Department of Anesthesiology and Intensive care for creating a friendly and supportive atmosphere and for all the help you have given me.

My new colleagues at the Department of Thoracic Surgery and Anesthesia, for kindly encouraging me in my work, and for creating an extremely im- pressive working environment.

Lucian Covaciu, for your support and friendship, and for being my friend when I needed it most.

My friends Silvia and Leif Nyholm who make my personal and family life richer.

Tibor Huzevka, for introducing me to Torsten Gordh.

My far-away family, sister Andrea and aunt Maria, for love that does not know distance.

And last, but not least, my wonderful and loving family: my wife Lubica, my son Samuel, and my daughter Emma. Lubica, for your strength and calm, for your endless support and quiet love, for being with me, and for just being. Samuel and Emma, for reminding me what is most important in my life, for the happiness you bring, for convincing me that heaven can exist on earth, and for all experiences you have given me, resulting in the strength to cope with things I previously thought were impossible.

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Acta Universitatis Upsaliensis Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine 1102 Editor: The Dean of the Faculty of Medicine

A doctoral dissertation from the Faculty of Medicine, Uppsala University, is usually a summary of a number of papers. A few copies of the complete dissertation are kept at major Swedish research libraries, while the summary alone is distributed internationally through the series Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine. (Prior to January, 2005, the series was published under the title “Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine”.)

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